EP0578784A1 - Odorant receptors ans uses thereof - Google Patents

Abstract

The present invention relates to an isolated nucleic acid, for example cDNA encoding a receptor for odorous substances, as well as to expression vectors containing such a nucleic acid, and to a purified protein encoding a receptor for odorous substances , with the above-mentioned expression vectors, and to the resulting transformed cell. Also described are methods of identifying ligands of odorous substances and identifying receptors for these substances, as well as methods of developing aromas, identifying compounds that suppress appetite and regulating appetite, as well as methods of controlling parasite colonies, stimulating and inhibiting fertility and detecting odors.

Description

ODORANT RECEPTORS AND USES THEREOF

Background of the Invention This application is a continuation-in-part of U.S. Serial No.681,880, filed April 5, 1991, the contents of which are hereby incorporated by refe5ence.

Throughout this application, various publications are referenced by Arabic numerals within parentheses. Full citations for these publications may be found at the end of the specification immediately preceding the claims. The disclosures of these publications in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art as known to those skilled therein as of the date of the invention described and claimed herein.

In vertebrate sensory systems, peripheral neurons respond to environmental stimuli and transmit these signals to higher sensory centers in the brain where they are processed to allow the discrimination of complex sensory information. The delineation of the peripheral mechanisms by which environmental stimuli are transduced into neural information can provide insight into the logic underlying sensory processing. Our understanding of color vision, for example, emerged only after the observation that the discrimination of hue results from the blending of information from only three classes of photoreceptors (1, 2, 3, 4). The basic logic underlying olfactory sensory perception, however, has remained elusive. Mammals possess an olfactory system of enormous discriminatory power (5, 6). Humans, for example, are thought to be capable of distinguishing among thousands of distinct odors. The specificity of odor recognition is emphasized by the observation that subtle alterations in the molecular structure of an odorant can lead to profound changes in perceived odor.

The detection of chemically distinct odorant presumably results from the association of odorous ligands with specific receptors on olfactory neurons which reside in a specialized epithelium in the nose. Since these receptors have not been identified, it has been difficult to determine how odor discrimination might be achieved. It is possible that olfaction, by analogy with color vision, involves only a few odor receptors, each capable of interaction with multiple odorant molecules. Alternatively, the sense of smell may involve a large number of distinct receptors each capable of associating with one or a small number of odorant. In either case, the brain must distinguish which receptors or which neurons have been activated to allow the discrimination between different odorant stimuli. Insight into the mechanisms underlying olfactory perception is likeLY to depend upon the isolation of the odorant receptors, and the characterization of their diversity, specificity, and patterns of expression.

The primary events in odor detection occur in a specialized olfactory neuroepithelium located in the posterior recesses of the nasal cavity. Three cell types dominate this epithelium (Figure 1A) : the olfactory sensory neuron, the sustentacular or supporting cell, and the basal cell which is a stem cell that generates olfactory neurons throughout life (7, 8). The olfactory sensory neuron is bipolar: a dendritic process extends to the mucosal surface where it gives rise to a number of specialized cilia which provide an extensive, receptive surface for the interaction of odors with olfactory sensory neurons. The olfactory neuron also gives rise to an axon which projects to the olfactory bulb of the brain, the first relay in the olfactory system. The axons of the olfactory bulb neurons, in turn, project to subcortical and cortical regions where higher level processing of olfactory information allows the discrimination of odors by the brain. The initial events in odor discrimination are thought to involve the association of odors with specific receptors on the cilia of olfactory neurons. Selective removal of the cilia results in the loss of olfactory response (9). Moreover, in fish, whose olfactory system senses amino acids as odors, the specific binding of amino acids to isolated cilia has been demonstrated (10, 11). The cilia are also the site of olfactory signal transduction. Exposure of isolated cilia from rat olfactory epithelium to numerous odorant leads to the rapid stimulation of adenylyl cyclase and elevations in cyclic AMP (an elevation in IP3 in response to one odorant has also been observed) (12, 13, 14, 15). The activation of adenylyl cyclase is dependent on the presence of GTP and is therefore likely to be mediated by receptor-coupled GTP binding proteins (G-proteins) (16). Elevations in cyclic AMP, in turn, are thought to elicit depolarization of olfactory neurons by direct activation of a cyclic nucleotide-gated, cation permeable channel (17, 18). This channel is opened upon binding of cyclic nucleotides to its cytoplasmic domain, and can therefore transduce changes in intracellular levels of cyclic AMP into alterations in the membrane potential.

These observations suggest a pathway for olfactory signal transduction (Figure 1B) in which the binding of odors to specific surface receptors activates specific G-proteins. The G-proteins then initiate a cascade of intracellular signalling events leading to the generation of an action potential which is propagated along the olfactory sensory axon to the brain. A number of neurotransmitter and hormone receptors which transduce intracellular signals by activation of specific G-proteins have been identified. Gene cloning has demonstrated that each of these receptors is a member of a large superfamily of surface receptors which traverse the membrane seven times (19, 20). The pathway of olfactory signal transduction (Figure 1B) predicts that the odorant receptors might also be members of this superfamily of receptor proteins. The detection of odors in the periphery is therefore likely to involve signalling mechanisms shared by other hormone or neurotransmitter systems, but the vast discriminatory power of the olfactory system will require higher order neural processing to permit the perception of individual odors. This invention address the problem of olfactory perception at a molecular level. Eighteen different members of an extremely large multigene family have been cloned and characterized which encodes seven transmembrane domain proteins whose expression is restricted to the olfactory epithelium. The members of this novel gene family encode the individual odorant receptors.

SUMMARY OF THE INVENTION

The invention provides an isolated nucleic acid, e.g. a DNA and cDNA molecule, encoding an odorant receptor. The invention further provides expression vectors containing such nucleic acid. Also provided by the invention is a purified protein encoding an odorant receptor. The invention further provides a method of transforming cells which comprises transfecting a suitable host cell with a suitable expression vector containing the nucleic acid encoding the odorant receptor.

The invention also provides methods of identifying odorant ligands and of identifying odorant receptors. The invention further provides methods of developing fragrances, of identifying appetite suppressant compounds, of controlling appetite. The invention also provides methods of controlling insect and other animal populations. The invention additionally provides a method of detecting odors such as the vapors emanating from Cocaine, Marijuana, Heroin, Hashish, Angel Dust, gasoline, decayed human flesh, alcohol, gun powder explosives, plastic explosives, firearms, poisonous or harmful smoke, or natural gas.

Description of the Figures

Figure 1. The Olfactory Neuroepithelium and a Pathway for Olfactory Signal Transduction. A. The Olfactory Neuroepithelium. The initial event in odor perception occurs in the nasal cavity in a specialized neuroepithelium which is diagrammed here. Odors are believed to interact with specific receptors on the cilia of olfactory sensory neurons. The signal generated by these initial binding events are propagated by olfactory neuron axons to the olfactory bulb. B. A Pathway of Olfactory Signal Transduction. In this scheme, the binding of an odorant molecule to an odor-specific transmembrane receptor leads to the interaction of the receptor with a GTP-binding protein (G_s[olf]). This interaction, in turn, leads to the release of the GTP-coupled α-subunit of the G-protein, which then stimulates adenylyl cyclase to produce elevated levels of cAMP. The increase in cAMP opens nucleotide-gated cation channels, thus causing an alteration in membrane potential.

Figure 2. A PCR Amplification Product Containing Multiple Species of DNA. cDNA prepared from olfactory epithelium RNA was subjected to PCR amplification with a series of different primer oligonucleotides and the DNA products of appropriate size were isolated, further amplified by PCR, and size fractionated on agarose gels (A) (For details, see text). Each of these semipurified PCR products was digested with the restriction enzyme, Hinf I, and analyzed by agarose gel electrophoresis. Lanes marked "M" contain size markers of 23.1, 9.4, 5.6, 4.4, 2.3, 2.0, 1.35, 1.08, 0.87, 0.60, 0.31, 0.28, 0.23, 0.19, 0.12 and 0.07kb. (B). Twenty-two of the 64 PCR products that were isolated and digested with Hinf I are shown here. Digestion of one of these, PCR 13, yielded a large number of fragments whose sizes summed to a value much greater than that of the undigested PCR 13 DNA, indicating that PCR 13 might contain multiple species of DNA which are representatives of a multigene family.

Figure 3. Northern Blot Analysis with a Mixture of Twenty Probes. One μg of polyA+ RNA isolated from rat olfactory epithelium, brain, or spleen was size-fractionated in formaldehyde agarose, blotted onto a nylon membrane, and hybridized with a ³²P-labeled mixture of segments of 20 cDNA clones. The DNA segments were obtained by PCR using primers homologous to transmembrane domains 2 and 7.

Figure 4. The Protein Sequences Encoded by Ten Divergent cDNA Clones. Ten divergent cDNA clones were subjected to DNA sequence analyses and the protein sequence encoded by each was determined. Amino acid residues which are conserved in 60% or more of the proteins are shaded. The presence of seven hydrophobic domains (I-VII), as well as short conserved motifs shared with other members of the superfamily, demonstrate that these proteins belong to the seven transmembrane domain protein superfamily. Motifs conserved among members of the superfamily and the family of olfactory proteins include the GN in TM1 (transmembrane domain 1), the central W of TM4 , the Y near the C-terminal end of TM5, and the NP in TM7. In addition, the DRY motif C-terminal to TM3 is common to many members of the G-protein-coupled superfamily. However, all of the proteins shown here share sequence motifs not found in other members of this superfamily and are clearly members of a novel family of proteins.

Figure 5. Positions of Greatest Variability in the Olfactory Protein Family. In this diagram, the protein encoded by cDNA clone 115 is shown traversing the plasma membrane seven times with its N-terminus located extracellularly, and its C-terminus intracellularly. The vertical cylinders delineate the seven putative α-helices spanning the membrane. Positions at which 60% or more of the 10 clones shown in Figure 4 share the same residue as 115 are shown as white balls. More variable residues are shown as black balls. The high degree of variability encountered in transmembrane domains III, IV, and V is evident in this schematic.

Figure 6. The Presence of Subfamilies in a Divergent Multigene Family. Partial nucleotide sequences and deduced protein sequences were obtained for 18 different cDNA clones. Transmembrane domain V along with the flanking loop sequences, including the entire cytoplasmic loop between transmembrane domains V and VI, are shown here for each protein. Amino acid residues found in 60% or more of the clones in a given position are shaded (A). This region of the olfactory proteins (particularly transmembrane domain V) appears to be highly variable (see Figure 4). These proteins, however, can be grouped into subfamilies (B,C,D) in which the individual subfamily members share considerable homology in this divergent region of the protein.

Figure 7. Southern Blot Analyses with Non-crosshvbridizing Fragments of Divergent cDNAs. Five μg of rat liver DNA was digested with Eco RI (A) or Hind III (B), electrophoresed in 0.75% agarose, blotted onto a nylon membrane, and hybridized to the ³²P-labeled probes indicated. The probes used were PCR-generated fragments of: 1, clone F9 (identical to F12 in Figure 4); 2, F5; 3, F6; 4, I3; 5, I7; 6, I14; or 7, I15. The lane labeled "1-7" was hybridized to a mixture of the seven probes. The probes used showed either no crosshybridization or only trace crosshybridization with one another. The size markers on the left correspond to the four blots on the left (1-4) whereas the marker positions noted on the right correspond to the four blots on the right ( 5-7 , " 1-7" ) .

Figure 8. Northern Blot Analysis with a Mix of Seven Divergent Clones. One μg of polyA+ RNA from each of the tissues shown was size-fractionated, blotted onto a nylon membrane, and hybridized with a ³²P-labeled mixture of segments of seven divergent cDNA clones (see Legend to Figure 7). Figure 9. The amino acid and nucleic acid sequence of clone

F3.

Figure 10. The amino acid and nucleic acid sequence of clone F5.

Figure 11. The amino acid and nucleic acid sequence of clone F6.

Figure 12. The amino acid and nucleic acid sequence of clone F12.

Figure 13. The amino acid and nucleic acid sequence of clone 13. Figure 14. The amino acid and nucleic acid sequence of clone 17.

Figure 15. The amino acid and nucleic acid sequence of clone 18.

Figure 16. The amino acid and nucleic acid sequence of clone 19.

Figure 17. The amino acid and nucleic acid sequence of clone 114. Figure 18. The amino acid and nucleic acid sequence of clone 115.

Figure 19. The amino acid and nucleic acid sequence of human clone H5.

Figure 20. The amino acid and nucleic acid sequence of clone J1, where the reading frame starts at nucleotide position 2.

Figure 21. The amino acid and nucleic acid sequence of clone J2.

Figure 22. The amino acid and nucleic acid sequence of clone J4, where the reading frame starts at nucleotide position 2.

Figure 23. The amino acid and nucleic acid sequence of clone J7, where the reading frame starts at nucleotide position 2.

Figure 24. The amino acid and nucleic acid sequence of clone J8, where the reading frame starts at nucleotide positon 2.

Figure 25. The amino acid and nucleic acid sequence of clone J11.

Figure 26. The amino acid and nucleic acid sequence of clone J14, where the reading frame starts at nucleotide position 2.

Figure 27. The amino acid and nucleic acid sequence of clone J15, where the reading frame starts at nucleotide psition 2. Figure 28. The amino acid and nucleic acid sequence of clone J16, where the reading frame starts at nucleotide position 2. Figure 29. The amino acid and nucleic acid sequence of clone J17, where the reading frame starts at nucleotide position 2.

Figure 30. The amino acid and nucleic acid sequence of clone J19, where the reading frame starts at nucleotide position 2.

Figure 31. The amino acid and nucleic acid sequence of clone J20, where the reading frame starts at nucleotide position 2.

Figure 32. SOUTHERN BLOT: Five micrograms of DNA isolated from 1. Human placenta, 2. NCI-H-1011 neuroblastoma cells, or 3. CHP 134 neuroblastoma cells were treated with the restriction enzyme A. Eco RI, B. Hind III, C. Bam HI, or D. Pst I, and then electrophoresed on an agarose gel and blotted onto a nylon membrane. The blotted DNA was hybridized to the 32P-labeled H3/H5 sequence. An autoradiograph of the hybridized blot is shown with the sizes of co-electrophoresed size markers noted in kilobases.

Detailed Description of the Invention

The invention provides an isolated nucleic acid, e.g. a DNA or cDNA molecule, encoding an odorant receptor. Such a receptor is a receptor which binds an odorant ligand and include but not limited to pheromone receptors. An odorant ligand may include, but is not limited to, molecules which interact with the olfactory sensory neuron, molecules which interact with the olfactory cilia, pheromones, and molecules which interact with structures within the vomeronasal organ.

The invention specifically provides the isolated cDNAs encoding odorant receptors the sequences of which are shown in Figures 9-31. The nucleic acid is most typically a cDNA and encodes an insect, a vertebrate, a fish or a mammalian odorant receptor. The mammalian odorant receptor is preferably a human, rat, mouse or dog receptor. In an embodiment, human odorant receptor cDNA sequence and the correspondent protein is isolated (Figure 19).

In another embodiment, phermone receptors are isolated and shown as clones J1, J2, J4, J7, J8, J11, J14, J15, J16, J17, J19 and J20 (Figures 20-31). The invention further provides expression vectors containing cDNA which encodes odorant receptors. Such expression vectors are well known in the art and include in addition to the nucleic acid the elements necessary for replication and expression in a suitable hosts. Suitable hosts are well known in the art and include without limitation bacterial hosts such as E. coli, animal hosts such as CHO cells, insect cells, yeast cells and like.

The invention also provides purified proteins encoding odorant receptors. Such proteins may be prepared by expression of the forementioned expression vectors in suitable host cells and recovery and purification of the receptors using methods well known in the art. Examples of such proteins include those having the amino acid sequences shown in figures 9-31.

The purified protein typically encodes an insect, vertebrate, fish or mammalian odorant receptor. The mammalian odorant receptor may be a human, rat, mouse or dog.

In one embodiment the invention provides a novel purified protein which belong to a class of proteins which have 7 transmembrane regions and a third cytoplasmic loop from the N-terminus which is approximately 17 amino acid long and to nucleic acid molecules encoding such proteins.

The invention provides methods of transforming cells which comprises transfecting a suitable host cell with a suitable expression vector containing nucleic acid encoding of the odorant receptor. Techniques for carrying out such transformations on cells are well known to those skilled in the art. (41,42) Additionally, the resulting transformed cells are also provided by the invention. These transformed cells may be either olfactory cells or non-olfactory cells. One advantage of using transformed non-olfactory cells is that the desired odorant receptor will be the only odorant receptor expressed on the cell's surface. In order to obtain cell lines that express a single receptor type, standard procedures may be used to clone individual cDNAs or genes into expression vectors and then transfect the cloned sequences into mammalian cell lines. This approach has been used with sequences encoding some other members of the seven transmembrane domain superfamily including the 5HT1c serotonin receptor. (43) The cited work illustrates how members of this superfamily transferred into cell lines may generate immortal cell lines that express high levels of the transfected receptor on the cell surface where it will bind ligand and that such abnormally expressed receptor molecules can transduce signals upon binding to ligand.

The invention also provides a method of identifying a desired odorant ligand which comprises contacting transformed non-olfactory cells expressing a known odorant receptor with a series of odorant ligands to determining which ligands bind to the receptors present on the non- olfactory cells.

Additionally, the invention provides a method of identifying a desired odorant receptor comprising contacting a series of transformed non-olfactory cells with a known odorant ligand and determining which odorant receptor binds with the odorant ligand.

The invention provides a method of detecting an odor which comprises: a) identifying a odorant receptor which binds the desired odorant ligand and; b) imbedding the receptor in a membrane such that when the odorant ligand binds to the receptor so identified a detectable signal is produced. In one embodiment of the invention the membrane used in this method is cellular, including a membrane of an olfactory cell or a synthetic membrane.

The ligand tested for may be the vapors emanating from Cocaine, Marijuana, Heroin, Hashish, Angel Dust, gasoline, decayed human flesh, alcohol, gun powder explosives, plastic explosives or firearms. In another embodiment the ligand tested for may be natural gas, a pheromone, toxic fumes, noxious fumes or dangerous fumes.

In one embodiment of the invention the detectable signal is a lightbulb lighting up, a buzzer buzzing, a bell ringing, a color change, phosphorescence, or radioactivity.

The invention further provides a method of quantifying the amount of an odorant ligand present in a sample which comprises utilizing the above-mentioned method for odor detection and then quantifying the amount of signal produced.

The invention further provides a method of developing fragrances which comprises identifying a desired odorant receptor by the above method, then contacting non-olfactory cells, which have been transfected with an expression vector containing nucleic acid encoding the desired odorant receptor such that the receptor is expressed upon the surface of the non-olfactory cell, with a series of compounds to determine which compound or compounds bind the receptor.

The invention provides to a method of identifying an "odorant fingerprint" which comprises contacting a series of cells, which have been transformed such that each express a known odorant receptor, with a desired sample and determining the type and quantity of the odorant ligands present in the sample. The invention provides a method of identifying odorant ligands which inhibit the activity of a desired odorant receptor which comprises contacting the desired odorant receptor with a series of compounds and determining which compounds inhibit the odorant ligand - odorant receptor interaction. The invention also provides for a method of identifying appetite suppressant compounds which comprises identifying odorant ligands by the method mentioned in the preceding paragraph wherein the desired odorant receptor is that which is associated with the perception of food. Additionally, the invention provides a method of controlling appetite in a subject which comprises contacting the olfactory epithelium of the subject with these odorant ligands. Further the invention provides a nasal spray, to control appetite comprising the compounds identified by the above method in a suitable carrier.

The invention provides a method of trapping odors which comprises contacting a membrane which contains multiples of the desired odorant receptor, with a sample such that the desired odorant ligand is absorbed by the binding of the odorant ligand to the odorant receptor. The invention also provides an odor trap employing this method. The invention also provides a method of controlling pest populations which comprises identifying odorant ligands by the method mentioned above which are alarm odorant ligands and spraying the desired area with the identified odorant ligands. Additionally, provided by the invention is a method of controlling a pest population which comprises identifying odorant ligands by the above mentioned method, which interfere with the interaction between the odorant ligands and the odorant receptors which are associated with fertility. In one embodiment the pest population is a population of insects or rodents, including mice and rats.

The invention also provides a method of promoting fertility which comprises identifying odorant ligands which interact with the odorant receptors associated with fertility by the above mentioned method. Further, the invention provides a method of inhibiting fertility which comprises employing the above mentioned method to identifying odorant ligands which inhibit the interaction between the odorant ligands and the odorant receptors associated with fertility.

This invention is illustrated in the Experimental Detail section which follow. These sections are set forth to aid in an understanding of the invention but are not intended to, and should not be construed to, limit in any way the invention as set forth in the claims which follow thereafter.

EXPERIMENTAL DETAILS

MATERIALS AND METHODS

Polymerase Chain Reaction RNA was prepared from the olfactory epithelia of Sprague Dawley rats according to Chirgwin et al. (40) or using RNAzol B (Cinna/Biotecx) and then treated with DNase I (0.1 unit/μg RNA) (Promega). In order to obtain cDNA, this RNA was incubated at 0.1 μg/μl with 5 μM random hexamers (Pharmacia) 1 mM each of dATP, dCTP, dGTP, TTP, and 2 units/μl RNase inhibitor (Promega) in 10 mM TrisCl (pH 8.3), 50 mM KCl, 2.5 mM MgCl₂, and 0.001% gelatin for 10 min. at 22°C, and then for a further 45 min. at 37°C following the addition of 20 u./μl of Moloney murine leukemia virus reverse transcriptase (BRL). After heating at 95°C for 3 min., cDNA prepared from 0.2 μg of RNA was used in each of a series of polymerase chain reactions (PCR) containing 10 mM TrisCl (pH 8.3), 50 mM KC1, 1.5 mM MgCl₂, 0.001% gelatin, 200 μM each of dATP, dCTP, dGTP, and TTP, 2.5 u. Taq polymerase (Perkin Elmer Cetus), and 2 μM of each PCR primer. PCR reactions were performed according to the following schedule: 96°C for 45 sec., 55°C for 4 min. (or 45°C for 2 min.), 72°C for 3 min. with 6 sec. extension per cycle for 48 cycles. The primers used for PCR were a series of degenerate oligonucleotides made according to the amino acid sequences found in transmembrane domain 2 and 7 of a variety of different members of the 7 transmembrane domain protein superfamily (19). The regions used correspond to amino acids number 60-70 and 286-295 of clone 115 (Figure 4). Each of five different 5' primers were used in PCR reactions with each of six different 3' primers. The 5' primers had the sequences:

C AC A C CT

A1, AATTGGATICTIGTIAATCTIGCIGTIGCIGCIGA;

C C CA A C C

A2, AATTATTTTCTIGTIAATCTIGCITTIGCIGA;

CCA CC A C

A3, AATTTITTTATIATITCICTIGCITGIGCIGA;

A T C T ACT C

A4, CGITTICTIATGTGTAACCTITGCTTTGCIGA;

C CT TG

A5, ACIGTITATATIACICATCTIACIATIGCIGA.

The 3' primers were:

TTA T CAG C C A

B1, CTGICGGTTCATIAAIACATAIATIATIGGGTT;

TG GA G G A A

B2, GATCGTTIAGACAACAATAIATIATIGGGTT;

A G G A

B3, TCIATGTTAAAIGTIGTATAIATIATIGGGTT;

T G G A A

B4, GCCTTIGTAAAIATIGCATAIAGGAAIGGGTT;

G AGA G G G A

B5, AAATCIGGGCTICGICAATAIATCAIIGGGTT;

CT CT G G G B6, GAIGAICCIACAAAAAAATAIATAAAIGGGTT.

An aliquot of each PCR reaction was analyzed by agarose gel electrophoresis and bands of interest were amplified further by performing PCR reactions on pipet tip (approx. 1 μl) plugs of the agarose gels containing those DNAs. Aliquots of these semi-purified PCR products were digested with the restriction enzymes Hae III or Hinf I and the digestion products were compared with the undigested DNAs on agarose gels.

Isolation and Analysis of cDNA Clones CDNA libraries were prepared according to standard procedures (41, 42) in the cloning vector, λZAP II (Stratagene) using poly A⁺ RNA prepared from Sprague Dawley rat epithelia (see above) or from an enriched population of olfactory neurons which had been obtained by a 'panning' procedure, using an antibody against the H blood group antigen (Chembiomed) found on a large percentage of rat olfactory neurons. In initial library screens, 8.5 X 10⁵ independent clones from the olfactory neuron library and 1.8 X 10⁶ clones from the olfactory epithelium library were screened (41) with a ³²P-labeled probe (prime-it, Stratagene) consisting of a pool of gel-isolated PCR products obtained using primers A4 and B6 (see above) in PCR reactions using as template, olfactory epithelium cDNA, rat liver DNA, or DNA prepared from the two cDNA libraries. In later library screens, a mixture of PCR products obtained from 20 cDNA clones with the A4 and B6 primers was used as probe ('P1' probe). In initial screens, phage clones were analyzed by PCR using primers A4 and B6 and those which showed the appropriate size species were purified. In later screens, all position clones were purified, but only those that could be amplified with the B6 primer and a primer specific for vector sequence were analyzed further. To obtain plasmids from the isolated phage clones, phagemid rescue was performed according to the instructions of the manufacturer of λZAP II (Stratagene). DNA sequence analysis was performed on plasmid DNAs using the Sequenase system (USB), initially with the A4 and B6 primers and later with oligonucleotide primers made according to sequences already obtained. Northern and Southern Blot Analyses

For Northern blots, poly A⁺ RNAs from various tissues were prepared as described above or purchased from Clontech. One μg of each RNA was size fractionated on formaldehyde agarose gels and blotted onto nylon membranes (41, 42). For Southern blots, genomic DNA prepared from Sprague Dawley rat liver was digested with the restriction enzymes Eco RI or Hind III, size fractionated on agarose gels and blotted onto nylon membranes (41, 42). The membranes were dried at 80°C, and then prehybridized in 0.5 M sodium phosphate buffer (pH 7.3) containing 1% bovine serum albumin and 4% sodium dodecyl sulfate. Hybridization was carried out in the same buffer at 65°-70°C for 14-20 hrs. with DNAs labeled with ³²P. For the first Northern blot shown, the 'P1' probe (see above under cDNA clone isolation) was used. For the second

Northern blot shown, a mix of PCR fragments from seven divergent cDNA clones was used. For Southern blots, the region indicated in clone 115 by amino acids 118 through 251 was amplified from a series of divergent cDNA clones using PCR. The primers used for these reactions had the sequences:

P1, ATGGCITATGATCGITATGTIGC, and P4, AAIAGIGAIACIATIGAIAGATGIGAICC These DNAs (or a DNA encompassing transmembrane domains 2 through 7 for clone F6) were labeled and tested for crosshybridization at 70°C. Those DNAs which did not show appreciable crosshybridization were hybridized individually, or as a pool to Southern blots at 70°C.

Rat Sequences used to obtain similar sequences expressed in Humans There are genes similar to the rat genes discussed above present in humans, these genes may be readily isolated by screening human gene libraries with the cloned rate sequences or by performing PCR experiments on human genomic DNA with primers homologous to the rat sequences. First, PCR experiments were performed with genomic DNA from rat, human, mouse, and several other species. When primers homologous to transmembrane domains 2 and 6 (the A4/B6 primer set used to isolate the original rat sequences) were used, DNA of the appropriate size was amplified from rat, human and mouse DNAs. When these primary PCR reactions were subsequently diluted and subjected to PCR using primers to internal sequences (P1 and P4 primers), smaller DNA species were amplified whose size was that seen when the same primers were used in PCR reactions with the cloned rat cDNAs. Similarly, when the secondary PCR was performed with one outer primer together with one inner primer (ie. A4/P4 or P1/B6), amplified DNAs were obtained whose sizes were also consistent with the amplification of genes similar in sequence and organization to the cloned rat cDNAs. Second, a mix of segments from 20 of the rat cDNAs ('P1" probe) was used to screen libraries constructed from human genomic DNAs. Hybridization under high or low stringency conditions reveals the presence of a large number of cloned human DNA segments that are homologous to the rat sequences. Finally, RNA from a human olfactory tumor (neuroesthesioma, NCI-H-1011) cell line has been examined for sequences homologous to those cloned in the rat. cDNA prepared from this RNA was subjected to PCR with the A4/B6 primer set and a DNA species of the appropriate size was seen. This DNA was subcloned and partially sequenced and clearly encodes a member of the olfactory protein family identified in the rat.

The inserted sequence in human clones H3/H5 was amplified by PCR with the A4/B6 primers, gel purified, and then labeled with 32P. The labeled DNA was then hybridized to restriction enzyme human placenta. Multiple hybridizing species were observed with each DNA (See Figure 32). This observation is consistent with the presence of a family of odorant receptor genes in the human genome.

The sequence of clone H5 is hereby shown in Figure 19. In addition, the translated protein sequence is shown in Figure 19.

In other to identify odorant receptors in other species, degenerated primer oligonucleotides homologous to conserved regions within the rat odorant receptor family may be used in PCR reactions with genomic DNA or with cDNA prepared from olfactory tissue RNA from those species.

RESULTS

Cloning the Gene Family A series of degenerate oligonucleotides were designated which could anneal to conserved regions of members of the superfamily of G-protein coupled seven transmembrane domain receptor genes. Five degenerate oliogonucleotides (A1-5; see Experimental Procedures) matching sequences within transmembrane domain 2, and six degenerate oligonucleotides (B1-6) matching transmembrane domain 7 were used in all combinations in PCR reactions to amplify homologous sequences in cDNA prepared from rat olfactory epithelium RNA. The amplification products of each PCR reaction were then analyzed by agarose gel electrophoresis. Multiple bands were observed with each of the primer combinations. The PCR products within the size range expected for this family of receptors (600 to 1300 bp) were subsequently picked and amplified further with the appropriate primer pair in order to isolate individual PCR bands. Sixty-four PCR bands isolated in this fashion revealed only one or a small number of bands upon agarose gel electrophoresis. Representatives of these isolated PCR products are shown in Figure 2A.

The isolated PCR products were digested with the ndonuclease, Hae III or Hinf I, which recognize four base restriction sites and cut DNA at frequent intervals. In most instances, digestion of the PCR product with Hinf I generated a set of fragments whose molecular weights sum to the size of the original DNA (Figure 2B). These PCR bands are therefore likely to each contain a single DNA species. In some cases, however, restriction digestion yielded a series of fragments whose molecular weights sum to a value greater than that of the original PCR product. The most dramatic example is shown in Figure 2 where the 710 bp, PCR 13 DNA, is cleaved by Hinf I to yield a very large number of restriction fragments whose sizes sum to a value five- to ten-fold greater than that of the original PCR product. These observations indicated that PCR product 13 consists of a number of different species of DNA, each of which could be amplified with the same pair of primer oligonucleotides. In addition, when PCR experiments similar to those described were performed using cDNA library DNAs as templates, a 710 bp PCR product was obtained with the PCR13 primer pair (A4/B6) with DNA from olfactory cDNA libraries, but not a glioma cDNA library. Moreover, digestion of one of this 710 bp product also revealed the presence of multiple DNA species. In other cases (see PCR product 20, for example), digestion yielded a series of restriction fragments whose molecular weights also sum to a size greater than the starting material. Further analysis, however, revealed that the original PCR product consisted of multiple bands of similar but different sizes.

In order to determine whether the multiple DNA species present in PCR 13 encode members of a family of seven transmembrane domain proteins, PCR 13 DNA was cloned into the plasmid vector Bluescript and five individual clones were subjected to DNA sequence analysis. Each of the five clones exhibited a different DNA sequence, but each encoded a protein which displayed conserved features of the superfamily of seven transmembrane domain receptor proteins. In addition, the proteins encoded by all five clones shared distinctive sequence motifs not found in other superfamily members indicating they were all members of a new family of receptors.

To obtain full-length cDNA clones, cDNA libraries prepared from olfactory epithelium RNA or from RNA of an enriched population of olfactory sensory neurons were screened. The probe used in these initial screens was a mixture of PCR 13

DNA as well as DNA obtained by amplication of rat genomic

DNA or DNA from two olfactory cDNA libraries with the same primers used to generate PCR 13 (A4 and B6 primers).

Hybridizing plaques were subjected to PCR amplification with the A4/B6 primer set and only those giving a PCR product of the appropriate size (approximately 710 bp) were purified.

The frequency of such positive clones in the enriched olfactory neuron cDNA library was approximately five times greater than the frequency in the olfactory epithelium cDNA library. The increased frequency of positive clones observed in the olfactory neuron library is comparable to the enrichment in olfactory neurons generally obtained in the purification procedure.

The original pair of primers used to amplify PCR 13 DNA were then used to amplify coding segments of 20 different cDNA clones. A mix of these PCR products were labeled and used as probe for further cDNA library screens. This mixed probe was also used in a Northern blot (Figure 3) to determine whether the expression of the gene family is restricted to the olfactory epithelium. The mixed probe detects two diffuse bands centered at 2 and 5 kb in RNA from olfactory epithelium; no hybridization can be detected in brain or spleen. (Later experiments which examined a larger number of tissue RNAs with a more restricted probe will be shown below.) Taken together, these data indicate the discovery of a novel multigene family encoding seven transmembrane domain proteins which are expressed in olfactory epithelium, and could be expressed predominantly or exclusively in olfactory neurons.

The Protein Sequences of Numerous. Olfactory-specific Members of the Seven Transmembrane Domain Superfamily

Numerous clones were obtained upon screening cDNA libraries constructed from olfactory epithelium and olfactory neuron RNA at high stringency. Partial DNA sequences were obtained from 36 clones; 18 of these cDNA clones are different, but all of them encode proteins which exhibit shared sequence motifs indicating that they are members of the family identified in PCR 13 DNA. A complete nucleotide sequence was determined for coding regions of ten of the most divergent clones (Figure 4). The deduced protein sequences of these cDNAs defines a new multigene family which shares sequence and structural properties with the superfamily of neurotransmitter and hormone receptors that traverse the membrane seven times. This novel family, however, exhibits features different from any other member of the receptor superfamily thus far identified.

Each of the ten sequences contains seven hydrophobic stretches (19-26 amino acids) that represent potential transmembrane domains. These domains constitute the regions of maximal sequence similarity to other members of the seven transmembrane domain superfamily (see legend to Figure 4). On the basis of structural homologies with rhodopsin and the β-adrenergic receptors, (19) it is likely that the amino termini of the olfactory proteins are located on the extracellular side of the plasma membrane and the carboxyl termini are located in the cytoplasm. In this scheme, three extracellular loops alternate with three intracellular loops to link the seven transmembrane domains (see Figure 5). Analysis of the sequences in figure 4 demonstrates that the olfactory proteins, like other members of the receptor superfamily, display no evidence of an N-terminal signal sequence. As in several other superfamily members, a potential N-linked glycosylation site is present in all ten proteins within the short N-terminal extracellular segment. Other structural features conserved with previously identified members of the superfamily included cysteine residues at fixed positions within the first and second extracellular loops that are thought to form a disulfide bond. Finally, many of the olfactory proteins reveal a conserved cysteine within the C-terminal domain which may serve as a palmitoylation site anchoring this domain to the membrane (21). These features, taken together with several short, conserved sequence motifs (see legend to Figure 4), clearly define this new family as a member of the superfamily of genes encoding the seven transmembrane domain receptors.

There are, however, important differences between the olfactory protein family and the other seven transmembrane domain proteins described previously and these differences may be relevant to proposed function of these proteins in odor recognition. Structure-function experiments involving in vitro mutagenesis suggest that adrenergic ligands interact with this class of receptor molecule by binding within the plane of the membrane (22, 20). Not surprisingly, small receptor families that bind the same class of ligands, such as the adrenergic and muscarinic acetylcholine receptor families exhibit maximum sequence conservation (often over 80%) within the transmembrane domains. In contrast, the family of receptors discussed in this application shows striking divergence within the third, fourth, and fifth transmembrane domains (Figure 4). The variability in the three central transmembrane domains is highlighted schematically in Figure 5. The divergence in potential ligand binding domains is consistent with the idea that the family of molecules cloned is capable of associating with a large number of odorant of diverse molecular structure.

Receptors which belong to the superfamily of seven transmembrane domain proteins interact with G-proteins to generate intracellular signals. In vitro mutagenesis experiments indicate that one site of association between receptor and G-protein resides within the third cytoplasmic loop (22, 23). The sequence of this cytoplasmic loop in 18 different clones we have characterized is shown in Figure 6A. This loop which is often quite long and of variable length in the receptor superfamily is relatively short (only 17 amino acids) and of fixed length in the 18 clones examined. Eleven of the 18 different clones exhibit the sequence motif K/R I V S S I (or a close relative) at the N- terminus of this loop. Two of the cDNA clones reveal a different H I T C/W A V motif at this site. If this short loop is a site of contact with G-proteins, it is possible that the conserved motifs may reflect sites of interaction with different G-proteins that activate different intracellular signalling systems in response to odors. In addition, the receptors cloned reveal several serine or threonine residues within the third cytoplasmic loop. By analogy with other G-protein coupled receptors, these residues may represent sites of phosphorylation for specific receptor kinases involved in desensitization. (24) Subfamilies within the Multigene Family

Figure 6A displays the sequences of the fifth transmembrane domain and the adjacent cytoplasmic loop encoded by L8 of the cDNA clones we have analyzed. As a group, the 18 sequences exhibit considerable divergence within this region. The multigene family, however, can be divided into subfamilies such that the members of a given subfamily share significant sequence conservation. In subfamily B, clones F12 and F13, for example, differ from one another at only four of 44 positions (91% identify), and clearly define a subfamily. Clones F5 and 111 (subfamily D) differ from F12 and F13 at 34-36 positions within this region and clearly define a separate subfamily. Thus, this olfactory-specific multigene family consists of highly divergent subfamilies. If these genes encode odor receptors, it is possible that members of the divergent subfamilies bind odorant of widely differing structural classes. Members of the individual subfamilies could therefore recognize more subtle differences between molecules which belong to the same structural class of molecules structures. The Size of the Multigene Family

Genomic Southern blotting experiments were preformed and genomic libraries were screened to obtain an estimate of the sizes of the multigene family and the member subfamilies encoding the putative odor receptors. DNAs extending from the 3' end of transmembrane domain 3 to the middle of transmembrane domain 6 were synthesized by PCR from DNA of seven of the divergent cDNA clones (Figure 4). In initial experiments, these DNAs were labeled and hybridized to each other to define conditions under which minimal crosshybridization would be observed among the individual clones. At 70°C, the seven DNAs showed no crosshybridization, or crosshybridized only very slightly. The trace levels of crosshybridization observed are not likely to be apparent upon genomic Southern blot analysis where the amounts of DNA are far lower than in the test cross. Probes derived from these seven DNAs were annealed under stringent conditions, either individually or as a group, to Southern blots of rat liver DNA digested with the restriction endonucleases Eco RI or Hind III (Figure 7). Examination of the Southern blots reveals that all but one of the cDNAs detects a relatively large, distinctive array of bands in genomic DNA. Clone 115 (probe 7), for example, detects about 17 bands with each restriction endonuclease, whereas clone F9 (probe 1) detects only about 5-7 bands with each enzyme. A single band is obtained with clone 17 (probe 5). PCR experiments using nested primers (TM2/TM7 primers followed by primers to internal seguences) and genomic DNA as template indicate that the coding regions of the members of this multigene family, like those of many members of the G-protein coupled superfamily, may not be interrupted by introns. This observation, together with the fact that most of the probes only encompasses 400 nucleotides suggests that each band observed in these experiments is likely to represent a different gene. These data suggest that the individual probes chosen are representatives of subfamilies which range in size from a single member to as many as 17 members. A total of about 70 individual bands were detected in this analysis which could represent the presence of at least 70 different genes. Although the DNA probes used in these blots did not crosshybridize appreciably with each other, it is possible that a given gene might hybridize to more than one probe, resulting in an overestimate of gene number. However, it is probable that the total number of bands only reflects a minimal estimate of gene number since it is unlikely that we have isolated representative cDNAs from all of the potential subfamilies and the hybridizations were performed under conditions of very high stringency.

A more accurate estimate of the size of the olfactory-specific gene family was obtained by screening rat genomic libraries. The mix of the seven divergent probes used in Southern blots, or the mix of 20 different probes used in our initial Northern blots (see Figure 3), were used as hybridization probes under high (65°C) or lowered (55°C) stringency conditions in these experiments. Nested PCR (see above) was used to verify that the clones giving a positive signal under low stringency annealing conditions were indeed members of this gene family. It is estimated from these studies that there are between 100 and 200 positive clones per haploid genome. The estimate of the size of the family obtain from screens of genomic libraries again represents a lower limit. Given the size of the multigene family, one might anticipate that many of these genes are linked such that a given genomic clone may contain multiple genes. Thus the data from Southern blotting and screens of genomic libraries indicate that the multigene family identified consists of one to several hundred member genes which can be divided into multiple subfamilies.

It should be noted that the cDNA probes isolated may not be representative of the full complement of subfamilies within the larger family of olfactory proteins. The isolation of cDNAs, for example, relies heavily on PCR with primers from transmembrane domains 2 and 7 and biases our clones for homology within these regions. Thus, estimates of gene number as well as subsequent estimates of RNA abundance should be considered as minimal.

Expression of the Members of this Multigene Family Additional Northern blot analyses were preformed to demonstrate that expression of the members of this gene family is restricted to the olfactory epithelium. (Figure 8) Northern blot analysis with a mixed probe consisting of the seven divergent cDNAs used above reveals two diffuse bands about 5 and 2 kb in length in olfactory epithelium RNA. This pattern is the same as that seen previously with the mix of 20 DNAs. No annealing is observed to RNA from the brain or retina or other, nonneural tissues, including lung, liver, spleen, and kidney.

An estimate of the level of expression of this family can be obtained from screens of cDNA libraries. The frequency of positive clones in cDNA libraries made from olfactory epithelium RNA suggests that the abundance of the RNAs in the epithelium is about one in 20,000. The frequency of positive clones is approximately five-fold higher in a cDNA library prepared from RNA from purified olfactory neurons (in which 75% of the cells are olfactory neurons). The increased frequency of positive clones obtained in the olfactory neuron cDNA library is comparable to the enrichment we obtain upon purification of olfactory neurons. These observations suggest that this multigene family is expressed largely, if not solely, in olfactory neurons and may not be expressed in other cell types within the epithelium. If each olfactory neuron contains 10⁵ mRNA molecules, from the frequency of positive clones we predict that each neuron contains only 25-30 transcripts derived from this gene family. Since the family of olfactory proteins consists of a minimum of a hundred genes, a given olfactory neuron could maximally express only a proportion of the many different family members. These values thus suggest that olfactory neurons will exhibit significant diversity at the level of expression of these olfactory proteins.

Identification of pheromone receptors in vomeronasal organ The vomeronasal organ (vomeronasal gland) is an accessory olfactory structure that is located near the nasal cavity. Like the olfactory epithelium of the nasal cavity, the olfactory epithelium of the vomeronasal organ contains olfactory sensory neurons. The vomeronasal organ is believed to play an important role in the sensing of pheromones in numerous species. Pheromones are believed to have profound effects on both physiological and behavioral aspects of reproduction. the identification of pheromone receptors would permit the identification of the pheromones themselves. It would also enable one to identify agonists or antagonists that would either mimic the pheromones or block the pheromone receptors from transducing pheromone signals. Such information would be important to the development of species specific pesticides and, conversely, to animal husbandry. The identification of pheromone receptors in human could ultimately lead to the development of contraceptives or to treatments for infertility in humans. It is likely that the identification of pheromone receptors in low mammals such as rodents would lead to the identification of similar receptors in human.

In order to identify potential pheromone receptors, we isolate RNA from the vomeronasal organs of female rats and prepared cDNA from this RNA. The cDNA was subjected to PCR with several different pairs of degenerate oligonucleotide primers that match sequences present in the rat odorant receptor family. The PCR products were subcloned and the nucleotide sequences of the subcloned DNAs were determined. Each of the subcloned DNAs encodes a protein that belongs to the odorant receptor family. The sequences of the following vomeronasal subclones are shown: J1, J2, J4, J7, J8, J11, J14, J15, J16, J17, J19, J20. In a few cases (J2, J4), the same sequence was amplified with two different primer pairs and the sequence shown is a composite of the two sequences. It is possible that one or more of these molecules, or closely related molecules, serve as pheromone receptors in the rat.

DISCUSSION

The mammalian olfactory system can recognize and discriminate a large number of odorous molecules. Perception in this system, as in other sensory systems, initially involves the recognition of external stimuli by primary sensory neurons. This sensory information is then transmitted to the brain where it is decoded to permit the discrimination of different odors. Elucidation of the logic underlying olfactory perception is likely to require the identification of the specific odorant receptors, the analysis of the extent of receptor diversity and receptor specificity, as well as an understanding of the pattern of receptor expression in the olfactory epithelium. The odorant receptors are thought to transduce intracellular signals by interacting with G-proteins which activate second messenger systems (12, 13, 14, 15). These proteins are clearly members of the family of G-protein coupled receptors which traverse the membrane seven times (19). The odorant receptors should be expressed specifically in the tissue in which odorant are recognized. The family of olfactory proteins cloned is expressed in the olfactory epithelium. Hybridizing RNA is not detected in brain or retina, or in a host of nonneural tissues. Moreover, expression of this gene family the epithelium may be restricted to olfactory neurons. The family of odorant receptors must be capable of interacting with extremely diverse molecular structures. The genes cloned are members of any extremely large multigene family which exhibit variability in regions thought to be important in ligand binding. The possibility that each member of this large family of seven transmembrane proteins is capable of interacting with only one or a small number of odorant provides a plausible mechanism to accommodate the diversity of odor perception. The properties of the gene family identified suggests that this family is likely to encode a large number of distinct odorant receptors. Size of the Multigene Family

The size of the receptor repertoire is likely to reflect the range of detectable odors and the degree of structural specificity exhibited by the individual receptors. It has been estimated that humans can identify over 10,000 structurally-distinct odorous ligands. However, this does not necessarily imply that humans possess an equally large repertoire of odorant receptors. For example, binding studies in lower vertebrates suggest that structurally-related odorant may activate the same receptor molecules. In fish which smell amino acids, the binding of alanine to isolated cilia can be competed by other small polar residues (threonine and serine), but not by the basic amino acids, lysine or arginine (11). These data suggest that individual receptors are capable of associating with several structurally-related ligands, albeit with different affinities. Stereochemical models of olfactory recognition in mammals (25) (based largely on psychophysical, rather than biophysical data) have suggested existence of several primary odor groups including camphoraceous, musky, peppermint, ethereal, pungent, and putrid. In such a model, each group would contain odorant with common molecular configurations which bind to common receptors and share similar odor qualities.

Screens of genomic libraries with mixed probes consisting of divergent family members detect approximately 100 to 200 positive clones per genome. The present estimate of at least 100 genes provides only a lower limit since it is likely that the probes used do not detect all of the possible subfamilies. Moreover, it is probable that many of these genes are linked such that a given genomic clone may contain multiple genes. It is therefore expected that the actual size of the gene family may be considerably higher and this family of putative odorant receptors could constitute one of the largest gene families in the genome.

The characterization of a large multigene family encoding putative odorant receptors suggests that the olfactory system utilizes a far greater number of receptors than the visual system. Color vision, for example, allows the discrimination of several hundred hues, but is accomplished by only three different photoreceptors (1, 2, 3 and 4). The photoreceptors each have different, but overlapping absorption spectra which cover the entire spectrum of visible wavelengths. Discrimination of color results from comparative processing of the information from these three classes of photoreceptors in the brain. Whereas three photoreceptors can absorb light across the entire visible spectrum, our data suggest that a small number of odorant receptors cannot recognize and discriminate the full spectrum of distinct molecular structures perceived by the mammalian olfactory system. Rather, olfactory perception probably employs an extremely large number of receptors each capable of recognizing a small number of odorous ligands.

Diversity within the Gene Family and the Specificity of Odor Recognition The olfactory proteins identified in this application are clearly members of the superfamily of receptors which traverse the membrane seven time. Analysis of the proteins encoded by the 18 distinct cDNAs we have cloned reveals structural features which may render this family particularly well suited for the detection of a diverse array of structurally distinct odorant. Experiments with other members of this class of receptors suggest that the ligand binds to its receptor within the plane of the membrane such that the ligand contacts many, if not all of the transmembrane helices. The family of olfactory proteins can be divided into several different subfamilies which exhibit significant sequence divergence within the transmembrane domains. Nonconservative changes are commonly observed within blocks of residues in transmembrane regions 3, 4, and 5 (Figures 4, 5, 6); these blocks could reflect the sites of direct contact with odorous ligands. Some members, for example, have acidic residues in transmembrane domain 3, which in other families are thought to be essential for binding aminergic ligands (20) while other members maintain hydrophobic residues at these positions. This divergence within transmembrane domains may reflect the fact that the members of the family of odorant receptors must associate with odorant of widely different molecular structures.

These observations suggest a model in which each of the individual subfamilies encode receptors which bind distinct structural classes of odorant. Within a given subfamily, however, the sequence differences are far less dramatic and are often restricted to a small number of residues. Thus, the members of a subfamily may recognize more subtle variations among odor molecules of a given structural class. At a practical level, individual subfamilies may recognize grossly different structures such that one subfamily may associate, for example, with the aromatic compound, benzene and its derivatives, whereas a second subfamily may recognize odorous, short chain, aliphatic molecules. Subtle variations in the structure of the receptors within, for example, the hypothetical benzene subfamily could facilitate the recognition and discrimination of various substituted derivatives such as toluene, xylene or phenol. It should be noted that such a model, unlike previous stereochemical models, does not necessarily predict that molecules with similar structures will have similar odors. The activation of distinct receptors with similar structures could elicit different odors, since perceived odor will depend upon higher order processing of primary sensory information.

Evolution of the Gene Family and the Generation of Diversity

Preliminary evidence from PCR analyses suggests that members of this family of olfactory proteins are conserved in lower vertebrates as well as invertebrates. This gene family presumably expanded over evolutionary time providing mammals with the ability to recognize an increasing diversity of odorant. Examination of the sequences of the family members cloned from mammals provides some insight into the evolution of this multigene family. Although the chromosomal loci encoding these genes has yet to be characterized, it is likely that at least some member genes will be tandemly arranged in a large cluster as is observed with other large multigene families. A tandem array of this sort provides a template for recombination events including unequal crossing over and gene conversion, that can lead to expansion and further diversification of the sort apparent among the family members we have cloned (26).

The multigene family encoding the olfactory proteins is large: all of the member genes clearly have a common ancestral origin, but have undergone considerable divergence such that individual genes encode proteins that share from 40-80% amino acid identity. Subfamilies are apparent with groups of genes sharing greater homology among themselves than with members of other subfamilies. Examination of the sequences of even the most divergent subfamilies, however, reveals a pattern in which several blocks of conserved residues are interspersed with variable regions. This segmental homology is conceptually similar to the organization of framework and hypervariable domains within the families of immunoglobulin and T cell receptor variable region sequences (27, 28). This analogy goes beyond structural organization and may extend to the function of these two gene families: each family consists of a large number of genes which have diversified over evolutionary time to accommodate the binding of a highly diverse array of ligands. The evolutionary mechanisms responsible for the diversification and maintenance of these large gene families may also be similar. It has been suggested that gene conversion has played a major role in the evolution of immunoglobulin and T cell receptor variable domains (29, 30 and 31). Analysis of the sequence of the putative olfactory receptors reveals at least one instance where a motif from a variable region of one subfamily is found imbedded in the otherwise divergent sequence of a second subfamily, suggesting that conversion has occurred. Such a mixing of motifs from one subfamily to another over evolutionary time would provide additional combinatorial possibilities leading to the generation of diversity. It should be noted, however, that the combinatorial joining of gene segments by DNA rearrangement during development, which is characteristic of immunoglobulin loci (27), is not a feature of the putative odor receptor gene family. No evidence for DNA rearrangement to generate the diversity of genes cloned has been observed. The entire coding region has been sequenced along with parts of the 5' and 3' untranslated regions of 10 different cDNA clones. The sequences of the coding regions are all different; no evidence has been obtained for constant regions that would suggest DNA rearrangement of the sort seen in the immune system. The observations indicate that the diversity olfactory proteins are coded by a large number of distinct gene sequences. Although it is unlikely from the data that DNA rearrangement is responsible for the generation of diversity among the putative odorant receptors, it remains possible that DNA rearrangements may be involved in the regulation of expression of this gene family. If each olfactory neuron expresses only one or a small number of genes, then a transcriptional control mechanism must be operative to choose which of the more than one hundred genes within the family will be expressed in a given neuron. Gene conversion from one of multiple silent loci into a single active locus, as observed for the trypanosome-variable surface glycoproteins (32), provides one attractive model. The gene conversion event could be stochastic, such that a given neuron could randomly express any one of several hundred receptor genes, or regulated (perhaps by positional information), such that a given neuron could only express one or a small number of predetermined receptor types. Alternatively, it is possible that positional information in the olfactory epithelium controls the expression of the family of olfactory receptors by more classical mechanisms that do not involve DNA rearrangement. What ever mechanisms will regulate the expression of receptor genes within this large, multigene family, these mechanisms must accommodate the requirement that olfactory neurons are regenerated every 30-60 days (8) and therefore the expression of the entire repertoire of receptors must be accomplished many times during the life of an organism.

Receptor Diversity and the Central Processing of Olfactory Information

The results suggest the existence of a large family of distinct odorant receptors. Individual members of this receptor family are likely to be expressed by only a small set of the total number of olfactory neurons. The primary sensory neurons within the olfactory epithelium will therefore exhibit significant diversity at the level of receptor expression. The question then emerges as to whether neurons expressing the same receptors are localized in the olfactory epithelium. Does the olfactory system employ a topographic map to discriminate among the numerous odorant? The spatial organization of distinct classes of olfactory sensory neurons, as defined by receptor expression, can now be determined by using the procedures of in situ hybridization and immunohistochemistry with probes specific for the individual receptor subtypes. This information should help to distinguish between different models that have been proposed to explain the coding of diverse odorant stimuli (33). In one model, sensory neurons that express a given receptor and respond to a given odorant may be localized within defined positions within the olfactory epithelium. This topographic arrangement would also be reflected in the projection of olfactory sensory axons into discrete regions (glomeruli) within the olfactory bulb. In this scheme, the central coding to permit the discrimination of discrete odorant would depend, in part, on the spatial segregation of different receptor populations. Attempts to discern the topographic localization of specific receptors at the level of the olfactory epithelium has led to conflicting results. In some studies, electrophysiological recordings have revealed differences in olfactory responses to distinct odorant in different regions of the olfactory epithelium (34, 35). However, these experiments have been difficult to interpret since the differences in response across the epithelium are often small and are not observed in all studies (36).

A second model argues that sensory neurons expressing distinct odorant receptors are randomly distributed in the epithelium but that neurons responsive to a given odorant project to restricted regions within the olfactory bulb. In this instance, the discrimination of odors would be a consequence of the position of second order neurons in the olfactory bulb, but would be independent of the site of origin of the afferent signals within the epithelium.

Mapping of the topographic projections of olfactory neurons has been performed by extracellular recordings from different regions of the bulb (37, 38) and by 2-deoxyglucose autoradiography to map regional activity after exposure to different odorant (39). These studies suggest that spatially-localized groups of bulbar neurons preferentially respond to different odorant. The existence of specific odorant receptors, randomly distributed through the olfactory epithelium, which converge on a common target within the olfactory bulb, would raise additional questions about the recognition mechanisms used to guide these distinct axonal subsets to their central targets. Other sensory systems also spatially segregate afferent input from primary sensory neurons. The spatial segregation of information employed, for example, by the visual and somatosensory systems, is used to define the location of the stimulus within the external environment as well as to indicate the quality of the stimulus. In contrast, olfactory processing does not extract spatial features of the odorant stimulus. Relieved of the necessity to encode information about the spatial localization of the sensory stimulus, it is possible that the olfactory system of mammals uses the spatial segregation of sensory input solely to encode the identity of the stimulus itself. The molecular identification of the genes likely to encode a large family of olfactory receptors should provide initial insights into the underlying logic of olfactory processing in the mammalian nervous system.

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Molecular Cloning: A Laboratory Manual (Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press). 42. Sambrook, J., Fritsch, E.F., and Maniatis, T. (1989).

Molecular Cloning: A Laboratory Manual, Second Edition (Cold Spring Harbor, New York, Cold Spring Harbor Laboratory Press). 43. Julius et al., (1988). Molecular Charatization of a Functional cDNA Encoding a Seritonin 1c Receptor. Science 241, 558-564.

SEQUENCE LISTING

(1) GENERAL INFORMATION:

(i) APPLICANT: Columbia Univeraity in the City of N.Y.,

The Trustees of

(ii) TITLE OF INVENTION: ODORANT RECEPTORS AND USES THEREOF

(iii) NUMBER OF SEQUENCES: 36

(iv) CORRESPONDENCE ADDRESS:

(A) ADDRESSES: COOPER & DUNHAM

(B) STREET: 30 Rockefeller Plaza

(C) CITY: New York

(D) STATE: New York

(E) COUNTRY: U.S.A.

(F) ZIP: 10112

(v) COMPUTER READABLE FORM:

(A) MEDIUM TYPE: Floppy disk

(B) COMPUTER: IBM PC compatible

(C) OPERATING SYSTEM: PC-DOS/MS-DOS

(D) SOFTWARE: Patentln Release #1.0, Version #1.25

(vii) PRIOR APPLICATION DATA:

(A) APPLICATION NUMBER: US 681,880

(B) FILING DATE: 05-APR-1991

(viii) ATTORNEY/AGENT INFORMATION:

(A) NAME: White, John P.

(B) REGISTRATION NUMBER: 28,678

(C) REFERENCE/DOCKET NUMBER: 38586

(ix) TELECOMMUNICATION INFORMATION:

(A) TELEPHONE: (212) 977-9550

(B) TELEFAX: (212) 664-0525

(C) TELEX: (212) 422523 COOP UI

(2) INFORMATION FOR SEQ ID NO:1:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 954 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE: (B) CLONE: F12

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:1:

ATGGAATCAG GG AACAGCAC AAGAAGATTT TCAAGTTTTT TTCTTCTTGG ATTTACAGAA 60

AACCCACAAC TTCACTTCCT CATTTTTGCA CTATTCCTGT CCATGTACCT GGTAACAGTG 120

CTTGGGAACC TGCTTATCAT TATGGCCATC ATCACACAGT CTCATTTGCA TACACCCATG 180

TACTTTTTCC TTGCTAACCT ATCCTTTGTG GACATCTGTT TCACCTCCAC CACCATCCCA 240

AAGATGTTGG TAAATATATA CACCCAGAGC AAGAGCATCA CCTATGAAGA CTGTATTAGC 300

CAGATGTGTG TCTTCTTGGT TTTCGCAGAA TTGGGCAACT TTCTCCTGGC TGTGATGGCC 360

TATGACCGAT ATGTGGCTAA CTGTCACCCA CTGTGTTACA CAGTCATTGT GAACCACCGG 420

CTCTGTATCC TGCTGCTTCT GCTGTCCTGG GTTATCAGCA TTTTCCATGC CTTCATACAG 480

AGCTTAATTG TGCTACAGTT GACCTTCTGT GGAGATGTGA AAATCCCTCA CTTCTTCTGT 540

GAACTTAATC AGCTGTCCCA ACTCACCTGT TCAGACAACT TTCCAAGTCA CCTCATAATG 600

AATCTTGTAC CTGTTATGTT GGCAGCCATT TCCTTCAGTG GCATCCTTTA CTCTTATTTC 660

AAGATAGTAT CCTCCATACA TTCTATCTCC ACAGTTCAGG GGAAGTACAA GGCATTTTCT 720

ACTTGTGCCT CTCACCTTTC CATTGTCTCC TTATTTTATA GTACAGGCCT CGGAGTGTAC 780

GTCAGTTCTG CTGTGGTCCA AAGCTCACAT TCTGCTGCAA GTGCTTCGGT CATGTATACT 840

GTGGTCACCC CCATGCTGAA CCCCTTCATT TATAGTCTAA GGAATAAAGA TGTGAAGAGA 900

GCTCTGGAAA GACTGTTAGA AGGAAACTGT AAAGTGCATC ATTGGACTGG ATGA 954 (2) INFORMATION FOR SEQ ID NO: 2:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 1002 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: F3

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:2: ATGGACTCAA GCAACAGGAC AAGAGTTTCA GAATTTCTTC TTCTTGGATT TGTAGAAAAC 60

AAAGACCTAC AACCCCTTAT TTATGGTCTT TTTCTCTCTA TGTACCTGGT TACTGTCATT 120

GGAAACATAT CCATTATTGT GGCTATCATT TCAGATCCCT GTCTGCACAC CCCCATGTAT 180

TTCTTCCTCT CTAACCTGTC CTTTGTGGAC ATCTGTTTCA TTTCAACCAC TGTTCCAAAG 240

ATGTTAGTGA ACATCCAGAC CCAAAACAAT GTCATCACCT ATGCAGGATG CATTACCCAG 300

ATATACTTTT TCTTGCTCTT TGTAGAATTG GACAACTTCT TGCTGACTAT CATGGCCTAT 360

GACCGTTACG TAGCCATCTG TCACCCCATG CACTACACAG TTATCATGAA CTACAAGCTC 420

TGTGGATTTC TGGTTCTGGT ATCTTGGATT GTAAGTGTTC TGCATGCCTT GTTTCAAAGC 480

TTGATGATGT TGGCGCTGCC CTTCTGCACA CATCTGGAAA TCCCACACTA CTTCTGTGAA 540

CCTAATCAGG TGATTCAACT CACCTGTTCT GATGCATTTC TTAATGATCT TGTGATATAT 600

TTTACACTTG TGCTGCTGGC TACTGTTCCT CTTGCTGGCA TCTTCTATTC TTACTTCAAG 660

ATAGTGTCCT CCATATGTGC TATATCGTCA GTTCATGGGA AGTACAAAGC ATTCTCCACC 720

TGTGCATCTC ACCTTTCAGT CGTGTCTTTA TTTTACTGCA CAGGACTAGG AGTGTACCTC 780

AGTTCTGCTG CAAACAACAG CTCACAGGCA AGTGCCACAG CCTCAGTCAT GTACACTGTA 840

GTTACCCCTA TGGTGAACCC TTTTATCTAT AGTCTTAGGA ATAAAGATGT TAAGAGTGTT 900

CTGAAAAAAA CTCTTTGTGA GGAAGTTATA AGGAGTCCAC CTTCCCTACT TCATTTCTTC 960

CTAGTGTTAT GTCATCTCCC TTGTTTTATT TTTTGTTATT AA 1002 (2) INFORMATION FOR SEQ ID NO: 3:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 942 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: F5

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 3:

ATGAGCAGCA CCAACCAGTC CAGTGTCACC GAGTTCCTCC TCCTGGGACT CTCCAGGCAG 60

CCCCAGCAGC AGCAGCTCCT CTTCCTGCTC TTCCTCATCA TGTACCTGGC CACTGTCCTG 120 GGAAACCTGC TCATCATCCT GGCTATTGGC ACAGACTCCC GCCTGCACAC CCCCATGTAC 180

TTCTTCCTCA GTAACCTGTC CTTTGTGGAT GTCTGCTTCT CCTCTACCAC TGTCCCTAAA 240

GTTCTGGCCA ACCATATACT TGGGAGTCAG GCCATTTCCT TCTCTGGGTG TCTCACCCAG 300

CTGTATTTTC TCGCTGTGTT TGGTAACATG GACAATTTCC TGCTGGCTGT GATGTCCTAT 360

GACCGATTTG TGGCCATATG CCACCCTTTA CACTACACAA CAAAGATGAC CCGTCAGCTC 420

TGTGTCCTGC TTGTTGTGGG GTCATGGGTT GTAGCCAACA TGAATTGTCT GTTGCACATA 480

CTGCTCATGG CTCGACTCTC CTTCTGTGCA GACAACATGA TCCCCCACTT CTTCTGTGAT 540

GGAACTCCCC TCCTGAAACT CTCCTGCTCA GACACACATC TCAATGAGCT GATGATTCTT 600

ACAGAGGGAG CTGTGGTCAT GGTCACCCCA TTTGTCTGCA TCCTCATCTC CTACATCCAC 660

ATCACCTGTG CTGTCCTCAG AGTCTCATCC CCCAGGGGAG GATGGAAATC CTTCTCCACC 720

TGTGGCTCCC ACCTGGCTGT GGTCTGCCTC TTCTATGGCA CCGTCATCGC TGTGTATTTC 780

AACCCATCAT CCTCTCACTT AGCTGGGAGG GACATGGCAG CTGCAGTGAT GTATGCAGTG 840

GTGACCCCAA TGCTGAACCC TTTCATCTAT AGCCTGAGGA ACAGCGACAT GAAAGCAGCT 900

TTAAGGAAAG TGCTCGCCAT GAGATTTCCA TCTAAGCAGT AA 942 ( 2 ) INFORMATION FOR SEQ ID NO: 4 :

( i ) SEQUENCE CHARACTERISTICS :

( A ) LENGTH : 936 base pairs

( B ) TYPE : nucleic acid

( C ) STRANDEDNESS : single

( D ) TOPOLOGY : linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: F6

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:4:

ATGGCTTGGA GTACTGGCCA GAACCTGTCC ACACCAGGAC CATTCATCTT GCTGGGCTTC 60

CCAGGGCCAA GGAGCATGCG CATTGGGCTC TTCCTGCTTT TCCTGGTCAT GTATCTGCTT 120

ACGGTAGTTG GAAACCTAGC CATCATCTCC CTGGTAGGTG CCCACAGATG CCTACAGACA 180

CCCATGTACT TCTTCCTCTG CAACCTCTCC TTCCTGGAGA TCTGGTTCAC CACAGCCTGC 240

GTACCCAAGA CCCTGGCCAC ATTTGCGCCT CGGGGTGGAG TCATTTCCTT GGCTGGCTGT 300 GCCACACAGA TGTACTTTGT CTTTTCTTTG GGCTGTACCG AGTACTTCCT GCTGGCTGTG 360

ATGGCTTATG ACCGCTACCT GGCCATCTGC CTGCCACTGC GCTATGGTGG CATCATGACT 420

CCTGGGCTGG CGATGCGGTT GGCCCTGGGA TCCTGGCTGT GTGGGTTTTC TGCAATCACA 480

GTTCCTGCTA CCCTCATTGC CCGCCTCTCT TTCTGTGGCT CACGTGTCAT CAACCACTTC 540

TTCTGTGACA TTTCGCCCTG GATAGTGCTT TCCTGCACCG ACACGCAGGT GGTGGAACTG 600

GTGTCCTTTG GCATTGCCTT CTGTGTTATT CTGGCCTCGT GTGGTATCAC ACTAGTCTCC 660

TATGCTTACA TCATCACTAC CATCATCAAG ATTCCCTCTG CCCGGGGCCG GCACCGCGCC 720

TTCTCAACCT GCTCATCCCA TCTCACTGTG GTGCTGATTT GGTATGGCTC CACCATCTTC 780

TTGCATGTGA GGACCTCGGT AGAGAGCTCC TTGGACCTCA CCAAAGCTAT CACAGTGCTC 840

AACACCATTG TCACACCTGT GCTGAACCCT TTCATATATA CTCTGAGGAA CAAGGATGTC 900

AAGGAAGCTC TGCGCAGGAC GGTGAAGGGG AAGTGA 936 (2) INFORMATION FOR SEQ ID NO: 5:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 939 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

( F) TISSUE TYPE : olfactory epithelium

( vii ) IMMEDIATE SOURCE :

( B ) CLONE : 114

( xi ) SEQUENCE DESCRIPTION : SEQ ID NO: 5 :

ATGACTGGAA ATAACCAAAC TTTGATCTTG GAGTTCCTCC TCCTGGGTCT GCCCATCCCA 60

TCAGAGTATC ATCTCCTGTT CTATGCCCTG TTCCTGGCCA TGTACCTCAC CATCATCCTG 120

GGAAACCTGC TAATCATTGT CCTTGTTCGA CTGGACTCTC ATCTCCACAT GCCCATGTAC 180

TTGTTTCTCA GCAACTTGTC CTTCTCTGAC CTCTGCTTTT CCTCTGTCAC AATGCCCAAA 240

TTGCTTCAGA ACATGCAGAG CCAAGTACCA TCTATATCCT ATACAGGCTG CCTGACACAG 300

CTGTACTTCT TTATGGTTTT TGGAGATATG GAGAGCTTCC TTCTTGTGGT CATGGCCTAT 360

GACCGCTATG TGGCCATTTG CTTTCCTTTG CGTTACACCA CCATCATGAG CACCAAGTTC 420

TGTGCTTCAC TAGTGCTACT TCTGTGGATG CTGACGATGA CCCATGCCCT GCTGCATACC 480 CTACTCATTG CTAGATTGTC TTTTTGTGAG AAGAATGTGA TTCTTCACTT TTTCTGTGAC 540

ATTTCTGCTC TTCTGAAGTT GTCCTGCTCA GACATTTATG TTAATGAGCT GATGATATAT 600

ATCTTGGGTG GACTCATCAT TATTATCCCA TTCCTATTAA TTGTTATGTC CTATGTTAGA 660

ATTTTCTTCT CCATTTTGAA GTTTCCATCT ATTCAGGACA TCTACAAGGT ATTCTCAACC 720

TGTGGTTCCC ATCTGTCTGT GGTGACCTTG TTTTATGGGA CAATTTTTGG TATCTACTTA 780

TGTCCATCAG GTAATAATTC TACTGTGAAG GAGATTGCCA TGGCTATGAT GTACACAGTG 840

GTGACTCCCA TGCTGAATCC CTTCATCTAC AGCCTGAGGA ACAGAGACAT GAAAAGGGCC 900

CTAATAAGAG TTATCTGCAC TAAGAAAATC TCTCTGTAA 939 (2) INFORMATION FOR SEQ ID NO: 6:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 945 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

( B ) CLONE : 115

( xi ) SEQUENCE DESCRIPTION : SEQ ID NO: 6 :

ATGACAGAAG AGAACCAAAC TGTGATCTCC CAGTTCCTTC TCCTTTTCCT GCCCATCCCC 60

TCAGAGCACC AGCACGTGTT CTACGCCCTG TTCCTGTCCA TGTACCTCAC CACTGTCCTG 120

GGGAACCTCA TCATCATCAT CCTCATTCAC CTGGACTCCC ATCTCCACAC ACCCATGTAC 180

TTGTTTCTCA GCAACTTGTC CTTCTCTGAT CTCTGCTTTT CCTCTGTTAC GATGCCCAAG 240

TTGTTGCAGA ACATGCAGAG CCAAGTTCCA TCCATCCCCT TTGCAGGCTG CCTGACACAA 300

TTATACTTTT ACCTGTATTT TGCAGACCTT GAGAGCTTCC TGCTTGTGGC CATGGCCTAT 360

GACCGCTATG TGGCCATCTG CTTCCCCCTT CATTACATGA GCATCATGAG CCCCAAGCTC 420

TGTGTGAGTC TGGTGGTGCT GTCCTGGGTG CTGACCACCT TCCATGCCAT GCTGCACACC 480

CTGCTCATGG CCAGATTGTC ATTCTGTGCG GACAATATGA TCCCCCACTT TTTCTGTGAT 540

ATATCTCCTT TATTGAAACT GTCCTGCTCT GACACGCATG TTAATGAGTT GGTGATATTT 600 GTCATGGGAG GGCTTGTTAT TGTCATTCCA TTTGTGCTCA TCATTGTATC TTATGCACGA 660

GTTGTCGCCT CCATTCTTAA AGTCCCTTCT GTCCGAGGCA TCCACAAGAT CTTCTCCACC 720

TGCGGCTCCC ATCTGTCTGT GGTGTCACTG TTCTATGGGA CAATCATTGG TCTCTACTTA 780

TGTCCGTCAG CTAATAACTC TACTGTGAAG GAGACTGTCA TGGCCATGAT GTACACAGTG 840

GTGACCCCCA TGCTGAACCC CTTCATCTAC AGCCTGAGGA ACAGAGACAT GAAAGAGGCA 900

CTGATAAGAG TCCTTTGTAA AAAGAAAATT ACCTTCTGTC TATGA 945 (2) INFORMATION FOR SEQ ID NO: 7:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 933 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: 13

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 7:

ATGAACAATC AAACTTTCAT CACCCAATTC CTTCTCCTGG GACTGCCCAT CCCTGAAGAA 60

CATCAGCACC TGTTCTATGC CTTGTTCCTG GTCATGTACC TCACCACCAT CTTGGGAAAC 120

TTGCTAATCA TTGTACTTGT TCAACTGGAC TCCCAGCTCC ACACACCTAT GTATTTGTTT 180

CTCAGCAATT TGTCTTTCTC TGATCTATGT TTTTCCTCTG TCACAATGCC CAAGCTGCTG 240

CAGAACATGA GGAGCCAGGA CACATCCATT CCCTATGGAG GCTGCCTGGC ACAAACATAC 300

TTCTTTATGG TTTTTGGAGA TATGGAGAGT TTCCTTCTTG TGGCCATGGC CTATGACCGC 360

TATGTGGCCA TCTGCTTCCC TCTGCATTAC ACCAGCATCA TGAGCCCCAA GCTCTGTACT 420

TGTCTAGTGC TGTTATTGTG GATGCTGACG ACATCCCATG CCATGATGCA CACACTGCTT 480

GCAGCAAGAT TGTCTTTTTG TGAGAACAAT GTGGTCCTCA ACTTCTTCTG TGACCTATTT 540

GTTCTCCTAA AGCTGGCCTG CTCAGACACT TATATTAATG AGTTGATGAT ATTTATCATG 600

AGTACACTCC TCATTATTAT TCCATTCTTC CTCATTGTTA TGTCCTATGC AAGGATCATA 660

TCCTCTATTC TTAAGGTTCC ATCTACCCAA GGCATCTGCA AGGTCTTCTC TACCTGTGGT 720 TCCCATCTGT CTGTAGTATC ACTGTTCTAT GGGACAATTA TTGGTCTCTA CTTATGTCCA 780

GCAGGTAATA ATTCCACTGT AAAAGAGATG GTCATGGCCA TGATGTACAC TGTGGTGACC 840

CCCATGCTGA ATCCCTTCAT CTACAGCCTA AGGAATAGAG ATATGAAGAG GGCCCTAATA 900

AGAGTTATCT GTAGTATGAA AATCACTCTG TAA 933 (2) INFORMATION FOR SEQ ID NO: 8:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 984 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

( F ) TISSUE TYPE : olfactory epithelium

( vii ) IMMEDIATE SOURCE :

( B ) CLONE : 17

( xi ) SEQUENCE DESCRIPTION : SEQ ID NO: 8 :

ATGGAGCGAA GGAACCACAG TGCGAGAGTG AGTGAATTTG TGTTGCTGGG TTTCCCAGCT 60

CCTGCCCCAC TGCGAGTACT ACTATTTTTC CTTTCTCTTC TGGACTATGT GTTGGTGTTG 120

ACTGAAAACA TGCTCATCAT TATAGCAATT AGGAACCACC CAACCCTCCA CAAACCCATG 180

TATTTTTTCT TGGCTAATAT GTCATTTCTG GAGATTTGGT ATGTCACTGT TACGATTCCT 240

AAGATGCTCG CTGGCTTCAT TGGTTCCAAG GAGAACCATG GACAGCTGAT CTCCTTTGAG 300

GCATGCATGA CACAACTCTA CTTTTTCCTG GGCTTGGGTT GCACAGAGTG TGTCCTTCTT 360

GCTGTGATGG CCTATGACCG CTATGTGGCT ATCTGTCATC CACTCCACTA CCCCGTCATT 420

GTCAGTAGCC GGCTATGTGT GCAGATGGCA GCTGGATCCT GGGCTGGAGG TTTTGGTATC 480

TCCATGGTTA AAGTTTTCCT TATTTCTCGC CTGTCTTACT GTGGCCCCAA CACCATCAAC 540

CACTTTTTCT GTGATGTGTC TCCATTGCTC AACCTGTCAT GCACTGACAT GTCCACAGCA 600

GAGCTTACAG ACTTTGTCCT GGCCATTTTT ATTCTGCTGG GACCGCTCTC TGTCACTGGG 660

GCATCCTACA TGGCCATCAC AGGTGCTGTG ATGCGCATCC CCTCAGCTGC TGGCCGCCAT 720

AAAGCCTTTT CAACCTGTGC CTCCCACCTC ACTGTTGTGA TCATCTTCTA TGCAGCCAGT 780

ATTTTCATCT ATGCCAGGCC TAAGGCACTC TCAGCTTTTG ACACCAACAA GCTGGTCTCT 840

GTACTCTACG CTGTCATTGT ACCGTTGTTC AATCCCATCA TCTACTGCTT GCGCAACCAA 900 GATGTCAAAA GAGCGCTACG TCGCACGCTG CACCTGGCCC AGGACCAGGA GGCCAATACC 960

AACAAAGGCA GCAAAATTGG TTAG 984

(2) INFORMATION FOR SEQ ID NO: 9:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 939 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: CDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: 18

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 9:

ATGAACAACA AAACTGTCAT CACCCATTTC CTCCTCCTGG GATTGCCCAT CCCCCCAGAG 60

CACCAGCAAC TGTTCTTTGC CCTGTTCCTG ATCATGTACC TCACCACCTT TCTGGGAAAC 120

CTGCTAATTG TTGTCCTTGT TCAACTGGAC TCTCATCTCC ACACACCCAT GTACTTGTTT 180

CTCAGCAACT TGTCCTTCTC TGATCTCTGC TTTTCCTCTG TTACAATGCT GAAATTGCTG 240

CAAAATATAC AGAGCCAAGT ACCATCTATA TCCTATGCAG GATGCCTGAC ACAGATATTC 300

TTCTTTTTGT TGTTTGGCTA CCTTGGGAAT TTCCTTCTTG TAGCCATGGC CTATGACCGC 360

TATGTGGCCA TCTGCTTCCC TCTGCATTAT ACCAACATCA TGAGCCATAA GCTCTGTACT 420

TGTCTCCTGC TGGTATTTTG GATAATGACA TCATCTCATG CCATGATGCA CACCCTGCTT 480

GCAGCAAGAT TGTCTTTTTG TGAGAACAAT GTACTCCTCA ACTTTTTCTG TGACCTGTTT 540

GTTCTCCTAA AGTTGGCCTG CTCAGACACT TATGTTAATG AGTTGATGAT ACATATCATG 600

GGCGTGATCA TCATTGTTAT TCCATTCGTG CTCATTGTTA TATCCTATGC CAAGATCATC 660

TCCTCCATTC TTAAGGTTCC ATCTACTCAA AGCATTCACA AGGTCTTCTC CACTTGTGGT 720

TCTCATCTCT CTGTGGTGTC TCTGTTCTAC GGGACAATTA TTGGTCTCTA TTTATGTCCA 780

TCAGGTGATA ATTTTAGTCT AAAGGGGTCT GCCATGGCTA TGATGTACAC AGTGGTAACT 840

CCAATGCTGA ACCCGTTCAT CTACAGCCTA AGAAACAGAG ACATGAAGCA GGCCCTAATA 900

AGAGTTACCT GTAGCAAGAA AATCTCTCTG CCATGGTAG 939 (2) INFORMATION FOR SEQ ID NO: 10: (i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 945 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: 19

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 10:

ATGACTAGAA GAAACCAAAC TGCCATCTCT CAGTTCTTCC TTCTGGGCCT GCCATTCCCC 60

CCAGAGTACC AACACCTGTT CTATGCCCTG TTCCTGGCCA TGTACCTCAC CACTCTCCTG 120

GGGAACCTCA TCATCATCAT CCTCATTCTA CTGGACTCCC ATCTCCACAC ACCCATGTAC 180

TTGTTTCTCA GCAATTTATC CTTTGCCGAC CTCTGTTTTT CCTCTGTCAC AATGCCCAAG 240

TTGTTGCAGA ACATGCAGAG CCAAGTTCCA TCCATCCCCT ATGCAGGGTG CCTGGCACAG 300

ATATACTTCT TTCTGTTTTT TGGAGACCTT GGAAACTTCC TGCTTGTGGC CATGGCCTAT 360

GACCGCTATG TGGCCATCTG CTTCCCCCTT CATTACATGA GCATCATGAG CCCCAAGCTC 420

TGTGTGAGTC TGGTGGTGCT GTCCTGGGTG CTGACTACCT TCCATGCCAT GCTGCACACC 480

CTGCTCATGG CCAGATTGTC ATTCTGTGAG GACAGTGTGA TCCCTCACTA TTTCTGTGAT 540

ATGTCTACTC TGCTGAAAGT GGCTTGTTCT GACACCCATG ATAATGAATT AGCAATATTT 600

ATCTTAGGGG GCCCTATAGT TGTACTACCT TTCCTTCTCA TCATTGTTTC TTATGCAAGA 660

ATTGTTTCCT CCATCTTCAA GGTCCCTTCT TCTCAAAGCA TCCATAAAGC CTTCTCCACC 720

TGTGGCTCCC ACCTGTCTGT GGTGTCACTG TTCTATGGGA CAGTCATTGG TCTCTACTTA 780

TGTCCTTCAG CTAATAACTC CACTGTGAAG GAGACTGTCA TGTCTTTGAT GTACACAATG 840

GTGACACCCA TGCTGAACCC CTTCATCTAC AGCCTAAGAA ACAGAGACAT AAAAGATGCA 900

TTAGAAAAAA TAATGTGCAA AAAGCAAATT CCCTCCTTTC TATGA 945 (2) INFORMATION FOR SEQ ID NO: 11:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 645 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear (ii) MOLECULE TYPE: CDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: horaosapien

(vii) IMMEDIATE SOURCE:

(B) CLONE: H5

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 1..645

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:11:

ATC TGT TTT GTG TCT ACC ACT GTC CCA AAG CAG CTG GTG AAC ATC CAG 48 Ile Cys Phe Val Ser Thr Thr Val Pro Lys Gln Leu Val Asn Ile Gln

1 5 10 15

ACA CAG AGC AGA GTC ATC ACC TAT GCA GAC TGC ATC ACC CAG ATG TGC 96 Thr Gln Ser Arg Val Ile Thr Tyr Ala Asp Cys Ile Thr Gln Met Cys

20 25 30

TTT TTT ATA CTC TTT GTA GTG TTG GAC AGC TTA CTC CTG ACT GTG ATG 144 Phe Phe Ile Leu Phe Val Val Leu Asp Ser Leu Leu Leu Thr Val Met

35 40 45

GCC TAT GAC CGG TTT GTG GCC ATC TGT CAC CCC CTG CAC TAC ACA GTC 192 Ala Tyr Asp Arg Phe Val Ala Ile Cys His Pro Leu His Tyr Thr Val

50 55 60

ATT ATG AGC TCC TGG CTC TGT GGA CTG CTG GTT CTG GTG TCC TGG ATC 240 Ile Met Ser Ser Trp Leu Cys Gly Leu Leu Val Leu Val Ser Trp Ile

65 70 75 80

GTG AGC ATC CTA TAT TCT CTG TTA CAA AGC ATA ATG GCA TTG CAG CTG 288 Val Ser Ile Leu Tyr Ser Leu Leu Gln Ser Ile Met Ala Leu Gln Leu

85 90 95

TCC TTC TGT ACA GAA CTG AAA ATC CCT CAA TTT TTC TGT GAA CTT AAT 336 Ser Phe Cys Thr Glu Leu Lys Ile Pro Gln Phe Phe Cys Glu Leu Asn

100 105 110

CAG GTC ATC CAC CTT GCC TGT TCC GAC ACT TTT ATT AAT GAC ATG ATG 384 Gln Val Ile His Leu Ala Cys Ser Asp Thr Phe Ile Asn Asp Met Met

115 120 125

ATG AAT TTT ACA AGT GTG CTG CTG GGT GGG GGA TGC CTC GCT GGA ATA 432 Met Asn Phe Thr Ser Val Leu Leu Gly Gly Gly Cys Leu Ala Gly Ile

130 135 140

TTT TAC TNN TAC TTT AAG ATA CTT TGT TGC ATA TGT TCG ATC TCA TCA 480 Phe Tyr Xaa Tyr Phe Lys Ile Leu Cys Cys Ile Cys Ser Ile Ser Ser

145 150 155 160

GCT CAG GGG ATG AAT AAA GCA CTT TCC ACC TGT GCA TCT CAC CTC TCA 528 Ala Gln Gly Met Asn Lys Ala Leu Ser Thr Cys Ala Ser His Leu Ser

165 170 175 GTT GTC TCC TTA TTT TAT TGT ACA GGC GTA GGT GTG TAC CTT AGT TCT 576 Val Val Ser Leu Phe Tyr Cys Thr Gly Val Gly Val Tyr Leu Ser Ser

180 185 190

GCT GCA ACC CAT AAC TCA CTC TCA AAT GCT GCA GCC TCG GTG ATG TAC 624 Ala Ala Thr His Asn Ser Leu Ser Asn Ala Ala Ala Ser Val Met Tyr

195 200 205

ACT GTG GTC ACC TCC ATG CTC 645

Thr Val Val Thr Ser Met Leu

210 215

(2) INFORMATION FOR SEQ ID NO: 12:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 215 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 12:

Ile Cys Phe Val Ser Thr Thr Val Pro Lys Gln Leu Val Asn Ile Gln

1 5 10 15

Thr Gln Ser Arg Val Ile Thr Tyr Ala Asp Cys Ile Thr Gln Met Cys

20 25 30

Phe Phe Ile Leu Phe Val Val Leu Asp Ser Leu Leu Leu Thr Val Met

35 40 45

Ala Tyr Asp Arg Phe Val Ala Ile Cys His Pro Leu His Tyr Thr Val

50 55 60

Ile Met Ser Ser Trp Leu Cys Gly Leu Leu Val Leu Val Ser Trp Ile

65 70 75 80

Val Ser Ile Leu Tyr Ser Leu Leu Gln Ser Ile Met Ala Leu Gln Leu

85 90 95

Ser Phe Cys Thr Glu Leu Lys Ile Pro Gln Phe Phe Cys Glu Leu Asn

100 105 110

Gln Val Ile His Leu Ala Cys Ser Asp Thr Phe Ile Asn Asp Met Met

115 120 125

Met Asn Phe Thr Ser Val Leu Leu Gly Gly Gly Cys Leu Ala Gly Ile

130 135 140

Phe Tyr Xaa Tyr Phe Lys Ile Leu Cys Cys Ile Cys Ser Ile Ser Ser

145 150 155 160

Ala Gln Gly Met Asn Lys Ala Leu Ser Thr Cys Ala Ser His Leu Ser

165 170 175

Val Val Ser Leu Phe Tyr Cys Thr Gly Val Gly Val Tyr Leu Ser Ser

180 185 190

Ala Ala Thr His Asn Ser Leu Ser Asn Ala Ala Ala Ser Val Met Tyr

195 200 205 Thr Val Val Thr Ser Met Leu

210 215

(2) INFORMATION FOR SEQ ID NO: 13:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 640 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J1

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 2..640

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 13:

C ATC TGC TTT ACT TCT GCT AGC ATC CCA AAG ATG CTA GTG AAT ATA 46 Ile Cys Phe Thr Ser Ala Ser Ile Pro Lye Met Leu Val Asn Ile

1 5 10 15

CAG ACG AAG AAC AAG GTG ATC ACC TAT GAA GGC TGC ATC TCC CAA GTA 94 Gln Thr Lys Asn Lye Val Ile Thr Tyr Glu Gly Cys Ile Ser Gln Val

20 25 30

TAC TTT TCA TAC TCT TTG GAG TTT TGG ACA ACT TTC TTC TCG ACT GTG 142 Tyr Phe Ser Tyr Ser Leu Glu Phe Trp Thr Thr Phe Phe Ser Thr Val

35 40 45

ATG GCC TAT GAC CGA TAT GTG GCC ATC TGT CAC CCA TCT NAC TAC ACA 190 Met Ala Tyr Asp Arg Tyr Val Ala Ile Cys His Pro Ser Xaa Tyr Thr

50 55 60

GGT CAT CAT GAA CCN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 238

Gly His His Glu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

65 70 75

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 286

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

80 85 90 95

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 334

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

100 105 110

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 382 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa 115 120 125

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NTT 430 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

130 135 140

TAT TCT TAC TCT AAG ATA GTT TCC TCC ATA CGA GAA ATC TCA TCA TCA 478 Tyr Ser Tyr Ser Lys Ile Val Ser Ser Ile Arg Glu Ile Ser Ser Ser

145 150 155

CAG GGA AAG TAC AAG NNA TTC TCC ACC TGT GCA TCC CAC CTC TCA GTT 526 Gln Gly Lys Tyr Lye Xaa Phe Ser Thr Cys Ala Ser His Leu Ser Val

160 165 170 175

GTT TCA TTA TTC TAT TCT ACA CTT TTG GGT GTG TAC CTT AGT TCT TCT 574 Val Ser Leu Phe Tyr Ser Thr Leu Leu Gly Val Tyr Leu Ser Ser Ser

180 185 190

TTT ACC CAA AAC TCA CAC TCA ACT GCA CGG GCA TCT GTT ATG TAC AGT 622 Phe Thr Gln Asn Ser His Ser Thr Ala Arg Ala Ser Val Met Tyr Ser

195 200 205

GTG GTC ACC CCC ATG TTG 640

Val Val Thr Pro Met Leu

210

(2) INFORMATION FOR SEQ ID NO: 14:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 213 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 14:

Ile Cys Phe Thr Ser Ala Ser Ile Pro Lys Met Leu Val Asn Ile Gln

1 5 10 15

Thr Lys Asn Lys Val Ile Thr Tyr Glu Gly Cys Ile Ser Gln Val Tyr

20 25 30

Phe Ser Tyr Ser Leu Glu Phe Trp Thr Thr Phe Phe Ser Thr Val Met

35 40 45

Ala Tyr Asp Arg Tyr Val Ala Ile Cys His Pro Ser Xaa Tyr Thr Gly

50 55 60

His His Glu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

65 70 75 80

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

85 90 95

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

100 105 110

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

115 120 125 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Tyr

130 135 140

Ser Tyr Ser Lys Ile Val Ser Ser Ile Arg Glu Ile Ser Ser Ser Gln

145 150 155 160

Gly Lys Tyr Lys Xaa Phe Ser Thr Cys Ala Ser His Leu Ser Val Val

165 170 175

Ser Leu Phe Tyr Ser Thr Leu Leu Gly Val Tyr Leu Ser Ser Ser Phe

180 185 190

Thr Gln Asn Ser His Ser Thr Ala Arg Ala Ser Val Met Tyr Ser Val

195 200 205

Val Thr Pro Met Leu

210

(2) INFORMATION FOR SEQ ID NO: 15:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 636 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J2

( ix) FEATURE :

(A) NAME/KEY: CDS

(B) LOCATION: 1..636

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 15:

ACC TCC ACC ACC ATC CCA AAG ATG CTG GTA AAT ATA CAC ACC CAG AGC 48 Thr Ser Thr Thr Ile Pro Lys Met Leu Val Asn Ile His Thr Gln Ser

1 5 10 15

AAT ACT ATC ACC TAT GAA GAC TGT ATT TCC CAG ATG TTT GTA CTC TTG 96 Asn Thr Ile Thr Tyr Glu Asp Cys Ile Ser Gln Met Phe Val Leu Leu

20 25 30

GTT TTT GGA GAA CTG GAC AAC TTT CTC CTG GCT GTG ATG GCC TAT GAT 144 Val Phe Gly Glu Leu Asp Asn Phe Leu Leu Ala Val Met Ala Tyr Asp

35 40 45

CGA TAT GTG GCT ATC TGT CAC CCA CTG TAT TAC ACA GTC ATT GTG AAC 192 Arg Tyr Val Ala Ile Cys His Pro Leu Tyr Tyr Thr Val Ile Val Asn

50 55 60 CAC CGA CTC TGT ATC CTG CTG CTT CTG CTG TCC TGG GTT GTC AGC ATT 240 His Arg Leu Cys Ile Leu Leu Leu Leu Leu Ser Trp Val Val Ser Ile

65 70 75 80

TTA CAT GCC TTC TTA CAG AGC TTA ATT GTA CTA CAG TTG ACC TTC TGT 288 Leu His Ala Phe Leu Gln Ser Leu Ile Val Leu Gln Leu Thr Phe Cys

85 90 95

GGA GAT GTG AAA ATC CCT CAC TTC TTC TGT GAG CTC AAT CAG CTG TCC 336 Gly Asp Val Lys Ile Pro His Phe Phe Cys Glu Leu Asn Gln Leu Ser

100 105 110

CAA CTC ACA TGT TCA GAC AAC TTT CCA AGT CAC CTC ACA ATG CAT CTT 384 Gln Leu Thr Cys Ser Asp Asn Phe Pro Ser His Leu Thr Met His Leu

115 120 125

GTA CCT GTT ATA TTT GCA GCT ATT TCC CTC AGT GGT ATC CTT TAC TCT 432 Val Pro Val Ile Phe Ala Ala Ile Ser Leu Ser Gly Ile Leu Tyr Ser

130 135 140

TAT TTC AAG ATA GTG TCT TCC ATA CGT TCT ATG TCC TCA GTT CAA GGG 480 Tyr Phe Lys Ile Val Ser Ser Ile Arg Ser Me. Ser Ser Val Gln Gly

145 150 155 160

AAG TAC AAG GCA TTT TCT ACA TGT GCC TCT CAC CTT TCC ATT GTC TCC 528 Lys Tyr Lys Ala Phe Ser Thr Cys Ala Ser His Leu Ser Ile Val Ser

165 170 175

TTA TTT TAT AGT ACA GGC CTC GGG GTG TAC GTC AGT TCT GCT GTG ATC 576 Leu Phe Tyr Ser Thr Gly Leu Gly Val Tyr Val Ser Ser Ala Val Ile

180 185 190

CGA AGC TCA CAC TCC TCT GCA AGT GCT TCG GTC ATG TAT ACT GTG GTC 624 Arg Ser Ser His Ser Ser Ala Ser Ala Ser Val Met Tyr Thr Val Val

195 200 205

ACC CCC ATG TTG 636

Thr Pro Met Leu

210

(2) INFORMATION FOR SEQ ID NO: 16:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 212 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 16:

Thr Ser Thr Thr Ile Pro Lys Met Leu Val Asn Ile His Thr Gln Ser

1 5 10 15

Asn Thr Ile Thr Tyr Glu Asp Cys Ile Ser Gln Met Phe Val Leu Leu

20 25 30

Val Phe Gly Glu Leu Asp Asn Phe Leu Leu Ala Val Met Ala Tyr Asp

35 40 45

Arg Tyr Val Ala Ile Cys His Pro Leu Tyr Tyr Thr Val Ile Val Asn 50 55 60

His Arg Leu Cys Ile Leu Leu Leu Leu Leu Ser Trp Val Val Ser Ile

65 70 75 80

Leu His Ala Phe Leu Gln Ser Leu Ile Val Leu Gln Leu Thr Phe Cys

85 90 95

Gly Asp Val Lys Ile Pro His Phe Phe Cys Glu Leu Asn Gln Leu Ser

100 105 110

Gln Leu Thr Cys Ser Asp Asn Phe Pro Ser His Leu Thr Met His Leu

115 120 125

Val Pro Val Ile Phe Ala Ala Ile Ser Leu Ser Gly Ile Leu Tyr Ser

130 135 140

Tyr Phe Lys Ile Val Ser Ser Ile Arg Ser Met Ser Ser Val Gln Gly

145 150 155 160

Lye Tyr Lye Ala Phe Ser Thr Cys Ala Ser His Leu Ser Ile Val Ser

165 170 175

Leu Phe Tyr Ser Thr Gly Leu Gly Val Tyr Val Ser Ser Ala Val Ile

180 185 190

Arg Ser Ser His Ser Ser Ala Ser Ala Ser Val Met Tyr Thr Val Val

195 200 205

Thr Pro Met Leu

210

(2) INFORMATION FOR SEQ ID NO: 17:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 646 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J4

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 2..646

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:17:

C ATA GGC TAT TCA TCT TCT GTC ACA CCC AAT ATG CTT GTC AAC TTC 46 Ile Gly Tyr Ser Ser Ser Val Thr Pro Asn Met Leu Val Asn Phe

1 5 10 15

CTT ATA AAG CAA AAT ACC ATC TCA TAC CTT GGA TGT TCT ATA CAG TTT 94 Leu Ile Lys Gln Asn Thr Ile Ser Tyr Leu Gly Cys Ser Ile Gln Phe

20 25 30

GGC TCA GCT GCT TTG TTT GGA GGT CTT GAA TGC TTC CTT CTG GCT GCC 142 Gly Ser Ala Ala Leu Phe Gly Gly Leu Glu Cys Phe Leu Leu Ala Ala

35 40 45

ATG GCG TAT GAT CGT TTT GTA GCA ATC TGC AAC CCA CTG CTT TAT TCA 190 Met Ala Tyr Asp Arg Phe Val Ala Ile Cys Asn Pro Leu Leu Tyr Ser

50 55 60

ACG AAA ATG TCC ACA CAA GTC TGT GTC CAG TTG GTT GTG GGA TCT TAT 238 Thr Lye Met Ser Thr Gln Val Cys Val Gln Leu Val Val Gly Ser Tyr

65 70 75

ATA GGG GGA TTT CTT AAT GCC TCC TCT TTT ACC CTT TCC TTT TTT TCC 286 Ile Gly Gly Phe Leu Asn Ala Ser Ser Phe Thr Leu Ser Phe Phe Ser

80 85 90 95

TTG TCC TTC TGT GGA CCA AAT AGA ATC AAT CAC TTT TAC TGT GAT TTT 334 Leu Ser Phe Cys Gly Pro Asn Arg Ile Asn His Phe Tyr Cys Asp Phe

100 105 110

GCT CCG TTA GTA GAA CTT TCT TGC TCT GAT GTC AGT GTT CCT GAT GCT 382 Ala Pro Leu Val Glu Leu Ser Cys Ser Asp Val Ser Val Pro Asp Ala

115 120 125

GTT ACC TCA TTT TCT GCT GCC TCA GTT ACT ATG CTC ACA GTG TTT ATC 430 Val Thr Ser Phe Ser Ala Ala Ser Val Thr Met Leu Thr Val Phe Ile

130 135 140

ATA GCC ATC TCC TAT ACC TAT ATC CTC ATC ACC ATC CTG AAG ATG CGT 478 Ile Ala Ile Ser Tyr Thr Tyr Ile Leu Ile Thr Ile Leu Lys Met Arg

145 150 155

TCC ACT GAG GGT CGA CAG AAA GCA TTC TCT ACC TGC ACT TCC CAC CTC 526 Ser Thr Glu Gly Arg Gln Lys Ala Phe Ser Thr Cys Thr Ser His Leu

160 165 170 175

ACT GCA GTC ACT CTG TGC TAT GGA ACC ATC ACA TTC ATC TAT GTG ATG 574 Thr Ala Val Thr Leu Cys Tyr Gly Thr Ile Thr Phe Ile Tyr Val Met

180 185 190

CCC AAG TCC AGC TAC TCC ACA GAC CAG AAC AAG GTG GTG TCT GTG TTT 622 Pro Lys Ser Ser Tyr Ser Thr Asp Gln Asn Lys Val Val Ser Val Phe

195 200 205

TAT ATG GTG GTG ATC CCC ATG TTG 646

Tyr Met Val Val Ile Pro Met Leu

210 215

(2) INFORMATION FOR SEQ ID NO: 18:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 215 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear ( ii ) MOLECULE TYPE : protein

( xi ) SEQUENCE DESCRIPTION : SEQ ID NO : 18 :

Ile Gly Tyr Ser Ser Ser Val Thr Pro Asn Met Leu Val Asn Phe Leu

1 5 10 15 Ile Lys Gln Asn Thr Ile Ser Tyr Leu Gly Cys Ser Ile Gln Phe Gly

20 25 30

Ser Ala Ala Leu Phe Gly Gly Leu Glu Cys Phe Leu Leu Ala Ala Met

35 40 45

Ala Tyr Asp Arg Phe Val Ala Ile Cys Asn Pro Leu Leu Tyr Ser Thr 50 55 60

Lys Met Ser Thr Gln Val Cys Val Gln Leu Val Val Gly Ser Tyr Ile 65 70 75 80

Gly Gly Phe Leu Asn Ala Ser Ser Phe Thr Leu Ser Phe Phe Ser Leu

85 90 95

Ser Phe Cys Gly Pro Asn Arg Ile Asn His Phe Tyr Cys Asp Phe Ala

100 105 110

Pro Leu Val Glu Leu Ser Cys Ser Asp Val Ser Val Pro Asp Ala Val

115 120 125

Thr Ser Phe Ser Ala Ala Ser Val Thr Met Leu Thr Val Phe Ile Ile 130 135 140

Ala Ile Ser Tyr Thr Tyr Ile Leu Ile Thr Ile Leu Lye Met Arg Ser 145 150 155 160

Thr Glu Gly Arg Gln Lye Ala Phe Ser Thr Cys Thr Ser His Leu Thr

165 170 175

Ala Val Thr Leu Cys Tyr Gly Thr Ile Thr Phe Ile Tyr Val Met Pro

180 185 190

Lye Ser Ser Tyr Ser Thr Asp Gln Asn Lys Val Val Ser Val Phe Tyr

195 200 205

Met Val Val Ile Pro Met Leu

210 215

(2) INFORMATION FOR SEQ ID NO: 19:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 481 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium (B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: o' factory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J7

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 2..481

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 19:

C ATC TGC AAG CCC CTG CAC TAC ACC ACC ATC ATG AAT AAC CGA GTG 46 Ile Cys Lye Pro Leu His Tyr Thr Thr Ile Met Asn Asn Arg Val

1 5 10 15

TGC ACA GTT CTA GTC CTC TCC TGT TGG TTT GCT GGC CTG TTG ATC ATC 94 Cys Thr Val Leu Val Leu Ser Cys Trp Phe Ala Gly Leu Leu Ile Ile

20 25 30

CTC CCA CCT CTT GGT CAT GGC CTC CAG CTG GAG TTC TGT GAC TCC AAT 142 Leu Pro Pro Leu Gly His Gly Leu Gln Leu Glu Phe Cys Asp Ser Asn

35 40 45

GTG ATT GAT CAT TTT GGC TGT GAT GCC TCT CCA ATT CTG CAG ATA ACC 190 Val Ile Asp His Phe Gly Cys Asp Ala Ser Pro Ile Leu Gln Ile Thr

50 55 60

TGC TCA GAC ACG GTA TTT ATA GAG AAA ATT GTC TTG GCT TTT GCC ATA 238 Cys Ser Asp Thr Val Phe Ile Glu Lye Ile Val Leu Ala Phe Ala Ile

65 70 75

CTG ACA CTC ATC ATT ACT CTG GTA TGT GTT GTT CTC TCC TAC ACA TAC 286 Leu Thr Leu Ile Ile Thr Leu Val Cys Val Val Leu Ser Tyr Thr Tyr

80 85 90 95

ATC ATC AAG ACC ATT TTA AAG TTT CCT TCT GCT CAA CAA AGA AAA AAG 334 Ile Ile Lye Thr Ile Leu Lys Phe Pro Ser Ala Gln Gln Arg Lye Lye

100 105 110

GCC TTT TCT ACA TGT TCT TCC CAC ATG ATT GTG GTT TCC ATC ACC TAT 382 Ala Phe Ser Thr Cys Ser Ser His Met Ile Val Val Ser Ile Thr Tyr

115 120 125

GGG AGC TGT ATT TTC ATC TAC ATC AAA CCT TCA GCG AAG GAA GGG GTA 430 Gly Ser Cys Ile Phe Ile Tyr Ile Lys Pro Ser Ala Lys Glu Gly Val

130 135 140

GCC ATC AAT AAG GTT GTA TCT GTG CTC ACA ACA TCA GTC GCC CCT TTG 478 Ala Ile Asn Lys Val Val Ser Val Leu Thr Thr Ser Val Ala Pro Leu

145 150 155

CTC 481

Leu

160

(2) INFORMATION FOR SEQ ID NO: 20:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 160 amino acids (B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:20:

Ile Cys Lye Pro Leu His Tyr Thr Thr Ile Met Asn Aen Arg Val Cys

1 5 10 15

Thr Val Leu Val Leu Ser Cys Trp Phe Ala Gly Leu Leu Ile Ile Leu

20 25 30

Pro Pro Leu Gly His Gly Leu Gln Leu Glu Phe Cys Aep Ser Asn Val

35 40 45

Ile Asp His Phe Gly Cys Asp Ala Ser Pro Ile Leu Gln Ile Thr Cys 50 55 60

Ser Asp Thr Val Phe Ile Glu Lys Ile Val Leu Ala Phe Ala Ile Leu 65 70 75 80

Thr Leu Ile Ile Thr Leu Val Cys Val Val Leu Ser Tyr Thr Tyr Ile

85 90 95 Ile Lye Thr Ile Leu Lys Phe Pro Ser Ala Gln Gln Arg Lys Lye Ala

100 105 110

Phe Ser Thr Cys Ser Ser His Met Ile Val Val Ser Ile Thr Tyr Gly

115 120 125

Ser Cys Ile Phe Ile Tyr Ile Lye Pro Ser Ala Lye Glu Gly Val Ala 130 135 140

Ile Asn Lye Val Val Ser Val Leu Thr Thr Ser Val Ala Pro Leu Leu 145 150 155 160

(2) INFORMATION FOR SEQ ID NO: 21:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 481 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J8

(ix) FEATURE:

(A) NAME/KEY: CDS ( B ) LOCATION : 2 . . 481 ( xi ) SEQUENCE DESCRIPTION : SEQ ID NO : 21 :

C ATC TGC CAC CC G CTC CAC TAC TCT CTT CTC ATG AGT CCT GAC AAC 46 Ile Cys His Pro Leu His Tyr Ser Leu Leu Met Ser Pro Asp Asn

1 5 10 15

TGT GCT GCT CTG GTA ACA GTC TCC TGG GTG ACA GGG GTG GGC ACG GGC 94 Cys Ala Ala Leu Val Thr Val Ser Trp Val Thr Gly Val Gly Thr Gly

20 25 30

TTC CTG CCT TCC CTC CTG ATT TCT AAG TTG GAC TTC TGT GGG CCC AAC 142 Phe Leu Pro Ser Leu Leu Ile Ser Lye Leu Asp Phe Cys Gly Pro Asn

35 40 45

CGC ATC AAC CAT TTC TTC TGT GAC CTC CCT CCA TTA ATC CAG CTG TCC 190 Arg Ile Asn His Phe Phe Cys Asp Leu Pro Pro Leu Ile Gln Leu Ser

50 55 60

TGC TCC AGC GTC TTT GTG ACA GAA ATG GCC ATC TTT GTC CTG TCC ATC 238 Cye Ser Ser Val Phe Val Thr Glu Met Ala Ile Phe Val Leu Ser Ile

65 70 75

GCT GTG CTC TGC ATC TGT TTC CTC CTA ACC CNN NNN TCC TAC ATT TTC 286 Ala Val Leu Cys Ile Cys Phe Leu Leu Thr Xaa Xaa Ser Tyr Ile Phe

80 85 90 95

ATA GTG TCC TCC ATT CTG AGA ATC CCT TCC ACT ACC GGC AGG ATG AAG 334 Ile Val Ser Ser Ile Leu Arg Ile Pro Ser Thr Thr Gly Arg Met Lye

100 105 110

ACA TTT TCT ACA TGT GGC TCC CAC CTG GCC GTG GTC ACC ATC TAC TAT 382 Thr Phe Ser Thr Cys Gly Ser His Leu Ala Val Val Thr Ile Tyr Tyr

115 120 125

GGG ACC ATG ATC TCC ATG TAT GTC GGC CCA AAT GCG CAT CTG TCC CCG 430 Gly Thr Met Ile Ser Met Tyr Val Gly Pro Asn Ala His Leu Ser Pro

130 135 140

GAG CTC AAC AAG GTC ATT TCT GTC TTC TAC ACT GTG ATC ACC CCA CTA 478

Glu Leu Asn Lye Val Ile Ser Val Phe Tyr Thr Val Ile Thr Pro Leu

145 150 155

CTG 481

Leu

160

(2) INFORMATION FOR SEQ ID NO: 22:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 160 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 22:

Ile Cys His Pro Leu His Tyr Ser Leu Leu Met Ser Pro Asp Asn Cys 1 5 10 15

Ala Ala Leu Val Thr Val Ser Trp Val Thr Gly Val Gly Thr Gly Phe

20 25 30

Leu Pro Ser Leu Leu Ile Ser Lye Leu Asp Phe Cys Gly Pro Asn Arg

35 40 45

Ile Asn His Phe Phe Cys Asp Leu Pro Pro Leu Ile Gln Leu Ser Cys

50 55 60

Ser Ser Val Phe Val Thr Glu Met Ala Ile Phe Val Leu Ser Ile Ala

65 70 75 80

Val Leu Cys Ile Cys Phe Leu Leu Thr Xaa Xaa Ser Tyr Ile Phe Ile

85 90 95

Val Ser Ser Ile Leu Arg Ile Pro Ser Thr Thr Gly Arg Met Lys Thr

100 105 110

Phe Ser Thr Cys Gly Ser His Leu Ala Val Val Thr Ile Tyr Tyr Gly

115 120 125

Thr Met Ile Ser Met Tyr Val Gly Pro Asn Ala His Leu Ser Pro Glu

130 135 140

Leu Asn Lys Val Ile Ser Val Phe Tyr Thr Val Ile Thr Pro Leu Leu

145 150 155 160

(2) INFORMATION FOR SEQ ID NO: 23:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 646 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J11

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 2..646

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 23:

N GTC TGC TTC TCC TCC ACC ACT GTC CCC AAG GTA CTG GCT AAC CAC 46

Val Cys Phe Ser Ser Thr Thr Val Pro Lys Val Leu Ala Asn His

1 5 10 15 ATA CTC AGT AGT CAG GCC ATT TCC TTC TCT GGG TGT CTA ACT CAG CTG 94 Ile Leu Ser Ser Gln Ala Ile Ser Phe Ser Gly Cys Leu Thr Gln Leu

20 25 30

TAT TTT CTC TGT GTG TCT GTG AAT ATG GAC AAT TTC CTG CTG GCT GTG 142 Tyr Phe Leu Cys Val Ser Val Asn Met Asp Asn Phe Leu Leu Ala Val

35 40 45

ATG GCC TAT GAC AGA TTT GTG GCC ATA TGC CAC CCT TTG TAC TAC ACA 190 Met Ala Tyr Asp Arg Phe Val Ala Ile Cys His Pro Leu Tyr Tyr Thr

50 55 60

ACA AAG ATG ACC CAC CAG CTC TGT GTC TTG CTG GTG TCT GGA TCA NNN 238 Thr Lys Met Thr His Gln Leu Cys Val Leu Leu Val Ser Gly ser Xaa

65 70 75

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 286 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa xaa

80 85 90 95

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 334 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

100 105 110

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 382 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

115 120 125

NNN NNN NNN NNN NNN NNN NNT GTG ATC ATG GTC ACC CCA TTT GTC TGC 430 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Val Ile Met Val Thr Pro Phe Val Cys

130 135 140

ATC CTC ATC TCT TAC ATC TAC ATC ACC AAT GCA GTC CTC AGA GTC TCA 478 Ile Leu Ile Ser Tyr Ile Tyr Ile Thr Asn Ala Val Leu Arg Val Ser

145 150 155

TCC TTT AGG GGA GGA TGG AAA GCC TTC TCC ACC TGT GGC TCA CAC CTG 526 Ser Phe Arg Gly Gly Trp Lys Ala Phe Ser Thr Cys Gly Ser His Leu

160 165 170 175

GCT GTG GTC TGC CTC TTC TAT GGC ACC ATC ATT GCT GTG TAT TTC AAT 574 Ala Val Val Cys Leu Phe Tyr Gly Thr Ile Ile Ala Val Tyr Phe Asn

180 185 190

CCT GTA TCT TCC CAT TCA TCT GAG AAG GAC ACT GCA GCA ACT GTG CTA 622 Pro Val Ser Ser His Ser Ser Glu Lys Asp Thr Ala Ala Thr Val Leu

195 200 205

TAC ACA GTG GTG ACT CCC ATG TTG 646

Tyr Thr Val Val Thr Pro Met Leu

210 215

(2) INFORMATION FOR SEQ ID NO: 24:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 215 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein ( xi ) SEQUENCE DESCRIPTION : SEQ ID NO: 24 :

Val Cys Phe Ser Ser Thr Thr Val Pro Lys Val Leu Ala Asn His Ile

1 5 10 15

Leu Ser Ser Gln Ala Ile Ser Phe Ser Gly Cys Leu Thr Gln Leu Tyr

20 25 30

Phe Leu Cys Val Ser Val Asn Met Asp Asn Phe Leu Leu Ala Val Met

35 40 45

Ala Tyr Asp Arg Phe Val Ala Ile Cys His Pro Leu Tyr Tyr Thr Thr 50 55 60

Lye Met Thr His Gln Leu Cys Val Leu Leu Val Ser Gly Ser Xaa Xaa 65 70 75 80

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

85 90 95

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

100 105 110

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

115 120 125

Xaa Xaa Xaa Xaa Xaa Xaa Val Ile Met Val Thr Pro Phe Val Cys Ile

130 135 140

Leu Ile Ser Tyr Ile Tyr Ile Thr λβn Ala Val Leu Arg Val Ser Ser 145 150 155 160

Phe Arg Gly Gly Trp Lys Ala Phe Ser Thr Cys Gly Ser His Leu Ala

165 170 175

Val Val Cys Leu Phe Tyr Gly Thr Ile Ile Ala Val Tyr Phe Aen Pro

180 185 190

Val Ser Ser His Ser Ser Glu Lye Asp Thr Ala Ala Thr Val Leu Tyr

195 200 205

Thr Val Val Thr Pro Met Leu

210 215

(2) INFORMATION FOR SEQ ID NO: 25:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 646 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium (vii) IMMEDIATE SOURCE:

(B) CLONE: J14

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 2..646

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:25:

T GTC TGC TTC TCC TCC ACC ACT GTC CCC AAG GTA CTG GCT AAC CAC 46

Val Cys Phe Ser Ser Thr Thr Val Pro Lys Val Leu Ala Asn His

1 5 10 15

ATA CTC AGT AGT CAG GCC ATT TCC TTC TCT GGG TGT CTA ACT CAG CTG 94 Ile Leu Ser Ser Gln Ala Ile Ser Phe Ser Gly Cys Leu Thr Gln Leu

20 25 30

TAT TTT CTC TGT GTG TCT GTG AAT ATG GAC AAT TTC CTG CTG GCT GTG 142 Tyr Phe Leu Cys Val Ser Val Asn Met Asp Aen Phe Leu Leu Ala Val

35 40 45

ATG GCC TAT GAC AGA TTT GTG GCC ATA TGC CAC CCT TTG TAC TAC ACA 190 Met Ala Tyr Asp Arg Phe Val Ala Ile Cys His Pro Leu Tyr Tyr Thr

50 55 60

ACA CCG ATG ACC CAC CAG CTC TGT GTC TTG CTG GTG TCT GGA TCA NNN 238 Thr Pro Met Thr His Gln Leu Cys Val Leu Leu Val Ser Gly Ser Xaa

65 70 75

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 286 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

80 85 90 95

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 334 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

100 105 110

NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN NNN 382 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

115 120 125

NNN NNN NNN NNN NNN NNN NNT GTG ATC ATG GTC ACC CCA TTT GTC TGC 430 Xaa Xaa Xaa Xaa Xaa Xaa Xaa Val Ile Met Val Thr Pro Phe Val Cys

130 135 140

ATC CTC ATC TCT TAC ATC TAC ATC ACC AAT GCA GTC CTC AGA GTC TCA 478 Ile Leu Ile Ser Tyr Ile Tyr Ile Thr Asn Ala Val Leu Arg Val Ser

145 150 155

TCC TTT AGG GGA GGA TGG AAA GCC TTC TCC ACC TGT GGC TCA CAC CTG 526 Ser Phe Arg Gly Gly Trp Lys Ala Phe Ser Thr Cys Gly Ser His Leu

160 165 170 175

GCT GTG GTC TGC CTC TTC TAT GGC ACC ATC ATT GCT GTG TAT TTC AAT 574 Ala Val Val Cys Leu Phe Tyr Gly Thr Ile Ile Ala Val Tyr Phe Asn

180 185 190

CCT GTA TCT TCC CAT TCA TCT GAG AAG GAC ACT GCA GCA ACT GTG CTA 622 Pro Val Ser Ser His Ser Ser Glu Lye Asp Thr Ala Ala Thr Val Leu

195 200 205 TAC ACA GTG GTG ACT CCC ATG TTG 646

Tyr Thr Val Val Thr Pro Met Leu

210 215

(2) INFORMATION FOR SEQ ID NO: 26:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 215 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 26:

Val Cys Phe Ser Ser Thr Thr Val Pro Lye Val Leu Ala Asn His Ile

1 5 10 15

Leu Ser Ser Gln Ala Ile Ser Phe Ser Gly Cys Leu Thr Gln Leu Tyr

20 25 30

Phe Leu Cye Val Ser Val Asn Met Asp Asn Phe Leu Leu Ala Val Met

35 40 45

Ala Tyr Asp Arg Phe Val Ala Ile Cys His Pro Leu Tyr Tyr Thr Thr

50 55 60

Pro Met Thr His Gln Leu Cys Val Leu Leu Val Ser Gly Ser Xaa Xaa

65 70 75 80

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

85 90 95

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

100 105 110

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

115 120 125

Xaa Xaa Xaa Xaa Xaa Xaa Val Ile Met Val Thr Pro Phe Val Cys Ile

130 135 140

Leu Ile Ser Tyr Ile Tyr Ile Thr Asn Ala Val Leu Arg Val Ser Ser

145 150 155 160

Phe Arg Gly Gly Trp Lye Ala Phe Ser Thr Cys Gly Ser His Leu Ala

165 170 175

Val Val Cye Leu Phe Tyr Gly Thr Ile Ile Ala Val Tyr Phe Aen Pro

180 185 190

Val Ser Ser His Ser Ser Glu Lys Asp Thr Ala Ala Thr Val Leu Tyr

195 200 205

Thr Val Val Thr Pro Met Leu

210 215

(2) INFORMATION FOR SEQ ID NO: 27:

(i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 481 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J15

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 2..481

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 27:

T ATC TGC AAC CCT CTG CGC TAC CCA GTG CTC ATG AGC GGC CGG GTG 46 Ile Cys Asn Pro Leu Arg Tyr Pro Val Leu Met Ser Gly Arg Val

1 5 10 15

TGC CTG CTC ATG GTC GTG GCC TCC TGG TTG GGA GGA TCC CTC AAC GCC 94 Cys Leu Leu Met Val Val Ala Ser Trp Leu Gly Gly Ser Leu Asn Ala

20 25 30

TCC ATT CAG ACT TCT CTG ACC CTT CAG TTC CCC TAC TGT GGA TCA CGG 142 Ser Ile Gln Thr Ser Leu Thr Leu Gln Phe Pro Tyr Cys Gly Ser Arg

35 40 45

AAG ATC TCC CAC TTC TTC TGT GAG GTG CCC TCG CTG CTG ANN NTG GCC 190 Lys Ile Ser His Phe Phe Cys Glu Val Pro Ser Leu Leu Xaa Xaa Ala

50 55 60

TGT GCA GAC ACT GAA GCC TAT GAG CAG GTA CTA TTT GTG ACA GGC GTG 238 Cys Ala Asp Thr Glu Ala Tyr Glu Gln Val Leu Phe Val Thr Gly Val

65 70 75

GTG GTC CTC CTG GTG CCC ATT ACA TTC ATT ACT GCC TCT TAT GCC CTC 286 Val Val Leu Leu Val Pro Ile Thr Phe Ile Thr Ala Ser Tyr Ala Leu

80 85 90 95

ATC CTG GCT GCT GTG CTC CGA ATG CAC TCT GCG GAG GGG AGT CAG AAG 334 Ile Leu Ala Ala Val Leu Arg Met His Ser Ala Glu Gly Ser Gln Lys

100 105 110

GCC CTA GCC ACA TGC TCC TCT CAC CTG ACA GTC GTC AAT CTC TTC TAT 382 Ala Leu Ala Thr Cys Ser Ser His Leu Thr Val Val Asn Leu Phe Tyr

115 120 125

GGG CCC CTT GTC TAC ACC TAC ATG TTA CCT GCT TCC TAT CAC TCA CCA 430 Gly Pro Leu Val Tyr Thr Tyr Met Leu Pro Ala Ser Tyr His Ser Pro

130 135 140 GGC CAA GAC GAC ATA GTA TCC GTC TTT TAC ACC GTT CTC ACA CCC ATG 478 Gly Gln Aep Asp Ile Val Ser Val Phe Tyr Thr Val Leu Thr Pro Met

145 150 155

CTT 481

Leu

160

(2) INFORMATION FOR SEQ ID NO: 28:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 160 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:28:

Ile Cys Asn Pro Leu Arg Tyr Pro Val Leu Met Ser Gly Arg Val Cys

1 5 10 15

Leu Leu Met Val Val Ala Ser Trp Leu Gly Gly Ser Leu Asn Ala Ser

20 25 30

Ile Gln Thr Ser Leu Thr Leu Gln Phe Pro Tyr Cys Gly Ser Arg Lye

35 40 45

Ile Ser His Phe Phe Cys Glu Val Pro Ser Leu Leu Xaa Xaa Ala Cys

50 55 60

Ala Asp Thr Glu Ala Tyr Glu Gln Val Leu Phe Val Thr Gly Val Val

65 70 75 80

Val Leu Leu Val Pro Ile Thr Phe Ile Thr Ala Ser Tyr Ala Leu Ile

85 90 95

Leu Ala Ala Val Leu Arg Met His Ser Ala Glu Gly Ser Gln Lye Ala

100 105 110

Leu Ala Thr Cys Ser Ser His Leu Thr Val Val Asn Leu Phe Tyr Gly

115 120 125

Pro Leu Val Tyr Thr Tyr Met Leu Pro Ala Ser Tyr His Ser Pro Gly

130 135 140

Gln Asp Asp Ile Val Ser Val Phe Tyr Thr Val Leu Thr Pro Met Leu

145 150 155 160

(2) INFORMATION FOR SEQ ID NO: 29:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 481 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES ( iv ) ANTI -SENSE : NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J16

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 2..481

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 29:

C ATC TGT AGG CCT CTT CAC TAT CCT ACC CTC ATG ACC CAG ACA CTG 46 Ile Cys Arg Pro Leu His Tyr Pro Thr Leu Met Thr Gln Thr Leu

1 5 10 15

TGT GCC AAG ATT GCC ACT GGT TGC TGG TTG GGA GGC TTG GCT GGG CCA 94 Cys Ala Lys Ile Ala Thr Gly Cys Trp Leu Gly Gly Leu Ala Gly Pro

20 25 30

GTG GTA GAA ATT TCC TTG GTG TCT CGT CTC CTT TTT TGT GGC CCC AAT 142 Val Val Glu Ile Ser Leu Val Ser Arg Leu Leu Phe Cys Gly Pro Aen

35 40 45

CAC ATT CAA CAC ATC TTT TGT GAT TTC CCA CCT GTG CTG AGC TTG GCT 190 His Ile Gln His Ile Phe Cys Asp Phe Pro Pro Val Leu Ser Leu Ala

50 55 60

TGT ACT GAT ACA TCA GTG AAT GTC CTG GTA GAT TTT ATT ATA AAC CTC 238 Cye Thr Aep Thr Ser Val Asn Val Leu Val Asp Phe Ile Ile Asn Leu

65 70 75

TGC AAG ATC CTG GCC ACC TTC CTG CTG ATC CTG AGC TCC TAC TTG CAG 286 Cys Lys Ile Leu Ala Thr Phe Leu Leu Ile Leu Ser Ser Tyr Leu Gln

80 85 90 95

ATA ATC CGC ACA GTG CTC AAG ATT CCT TCA GCT GCA GGC AAG AAG AAA 334 Ile Ile Arg Thr Val Leu Lys Ile Pro Ser Ala Ala Gly Lys Lys Lys

100 105 110

GCA TTC TCG ACT TGT GCC TCC CAT CTC ACT GTG GTT CTC ATC TTC TAT 382 Ala Phe Ser Thr Cys Ala Ser His Leu Thr Val Val Leu Ile Phe Tyr

115 120 125

GGG AGC ATC CTT TTC ATG TAT GTG CGG CTG AAG AAG ACT TAC TCC CTT 430 Gly Ser Ile Leu Phe Met Tyr Val Arg Leu Lye Lye Thr Tyr Ser Leu

130 135 140

GAC TAC GAC AGA GCC TTG GCA GTA GTC TAC TCC GTG GTT ACC CCT TTC 478 Asp Tyr Asp Arg Ala Leu Ala Val Val Tyr Ser Val Val Thr Pro Phe

145 150 155

CTG 481

Leu

160

(2) INFORMATION FOR SEQ ID NO: 30: (i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 160 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 30:

Ile Cys Arg Pro Leu His Tyr Pro Thr Leu Met Thr Gln Thr Leu Cys 1 5 10 15

Ala Lye Ile Ala Thr Gly Cye Trp Leu Gly Gly Leu Ala Gly Pro Val

20 25 30

Val Glu Ile Ser Leu Val Ser Arg Leu Leu Phe Cys Gly Pro Asn His

35 40 45

Ile Gln His Ile Phe Cys Asp Phe Pro Pro Val Leu Ser Leu Ala Cys 50 55 60

Thr Asp Thr Ser Val Asn Val Leu Val Asp Phe Ile Ile Asn Leu Cys 65 70 75 80

Lye Ile Leu Ala Thr Phe Leu Leu Ile Leu Ser Ser Tyr Leu Gln Ile

85 90 95 Ile Arg Thr Val Leu Lye Ile Pro Ser Ala Ala Gly Lye Lye Lye Ala

100 105 110

Phe Ser Thr Cys Ala Ser His Leu Thr Val Val Leu Ile Phe Tyr Gly

115 120 125

Ser Ile Leu Phe Met Tyr Val Arg Leu Lye Lye Thr Tyr Ser Leu Aep 130 135 140

Tyr Aep Arg Ala Leu Ala Val Val Tyr Ser Val Val Thr Pro Phe Leu

145 150 155 160

(2) INFORMATION FOR SEQ ID NO: 31:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 481 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J17 ( ix ) FEATURE :

(A) NAME/KEY: CDS

(B) LOCATION: 2..481

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:31:

A ATC TGC AAC CCA CTG CTT TAT TCC ACC AAA ATG TCC ACA CAA GTC 46 Ile Cys Asn Pro Leu Leu Tyr Ser Thr Lye Met Ser Thr Gln Val

1 5 10 15

TGT ATC CAG TTG GTT GCA GGA TCT TAT ATA GGG GGT TTT CTT AAT ACT 94 Cye Ile Gln Leu Val Ala Gly Ser Tyr Ile Gly Gly Phe Leu Asn Thr

20 25 30

TGC CTC ATC ATG TTT TAC TTT TTC TCT TTT CTC TTC TGT GGG CCA AAT 142 Cys Leu Ile Met Phe Tyr Phe Phe Ser Phe Leu Phe Cys Gly Pro Asn

35 40 45

ATA GTT GAT CAT TTT TTC TGT GAT TTT GCT CCT TTN NTG GAA CTT TCG 190 Ile Val Asp His Phe Phe Cys Asp Phe Ala Pro Xaa Xaa Glu Leu Ser

50 55 60

TGC TCT GAT GTG AGT GTC TCT GTA GTT GTT ATG TCA TTT TCT GCT GGC 238 Cys Ser Asp Val Ser Val Ser Val Val Val Met Ser Phe Ser Ala Gly

65 70 75

TCA GTT ACT ATG ATC ACA GTG TTT ATC ATA GCC ATC TCC TAT TCT TAC 286 Ser Val Thr Met Ile Thr Val Phe Ile Ile Ala Ile Ser Tyr Ser Tyr

80 85 90 95

ATC CTC ATC ACC ATC CTG AAG ATG TCC TCA ACT GAG GGC CGT CAC AAG 334 Ile Leu Ile Thr Ile Leu Lys Met Ser Ser Thr Glu Gly Arg His Lys

100 105 110

GCT TTC TCC ACA TGT ACC TCC CAC CTC ACT GCA GTC ACT CTC TAC TAT 382 Ala Phe Ser Thr Cys Thr Ser His Leu Thr Ala Val Thr Leu Tyr Tyr

115 120 125

GGC ACC ATT ACC TTC ATT TAT GTG ATG CCC AAG TCC ACA TAC TCT ACA 430 Gly Thr Ile Thr Phe Ile Tyr Val Met Pro Lys Ser Thr Tyr Ser Thr

130 135 140

GAC CAG AAC AAG GTG GTG TCT GTG TTT TAC ATG GTG GTG ATC CCA ATG 478 Asp Gln Asn Lye Val Val Ser Val Phe Tyr Met Val Val Ile Pro Met

145 150 155

TTG 481

Leu

160

(2) INFORMATION FOR SEQ ID NO: 32:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 160 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 32: Ile Cys Asn Pro Leu Leu Tyr Ser Thr Lye Met Ser Thr Gln Val Cys

1 5 10 15

Ile Gln Leu Val Ala Gly Ser Tyr Ile Gly Gly Phe Leu Asn Thr Cye

20 25 30

Leu Ile Met Phe Tyr Phe Phe Ser Phe Leu Phe Cys Gly Pro Asn Ile

35 40 45

Val Asp His Phe Phe Cys Asp Phe Ala Pro Xaa Xaa Glu Leu Ser Cys

50 55 60

Ser Asp Val Ser Val Ser Val Val Val Met Ser Phe Ser Ala Gly Ser

65 70 75 80

Val Thr Met Ile Thr Val Phe Ile Ile Ala Ile Ser Tyr Ser Tyr Ile

85 90 95

Leu Ile Thr Ile Leu Lye Met Ser Ser Thr Glu Gly Arg His Lye Ala

100 105 110

Phe Ser Thr Cys Thr Ser His Leu Thr Ala Val Thr Leu Tyr Tyr Gly

115 120 125

Thr Ile Thr Phe Ile Tyr Val Met Pro Lye Ser Thr Tyr Ser Thr Asp

130 135 140

Gln Asn Lye Val Val Ser Val Phe Tyr Met Val Val Ile Pro Met Leu

145 150 155 160

(2) INFORMATION FOR SEQ ID NO: 33:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 479 base pairs

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: cDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J19

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 2..479

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:33:

T ATC TGC CAC CCT CTG AAG TAC ACA GTT ATC ATG AAT CAC TAT TTT 46 Ile Cys His Pro Leu Lys Tyr Thr Val Ile Met Asn His Tyr Phe 1 5 10 15

TGT GTG ATG CTG CTG CTC TTC TCT GTG TTC GTT AGC ATT GCA CAT GCG 94 Cys Val Met Leu Leu Leu Phe Ser Val Phe Val Ser Ile Ala His Ala

20 25 30

TTG TTC CAC ATT TTA ATG GTG TTG ATA CTG ACT TTC AGC ACA AAA ACT 142 Leu Phe His Ile Leu Met Val Leu Ile Leu Thr Phe Ser Thr Lye Thr

35 40 45

GAA ATC CCT CAC TTT TTC TGT GAG CTG GCT CAT ATC ATC AAA CTT ACC 190 Glu Ile Pro His Phe Phe Cye Glu Leu Ala His Ile Ile Lye Leu Thr

50 55 60

TGT TCC GAT AAT TTT ATC AAC TAT CTG CTG ATA TAC ACA GAG TCT GTC 238 Cys Ser Asp Asn Phe Ile Asn Tyr Leu Leu Ile Tyr Thr Glu Ser Val

65 70 75

TTA TTT TTT GGT GTT CAT ATT GTA GGG ATC ATT TTG TCT TAT ATT TAC 286 Leu Phe Phe Gly Val His Ile Val Gly Ile Ile Leu Ser Tyr Ile Tyr

80 85 90 95

ACT GTA TCC TCA GTT TTA AGA ATG TCA TTA TTG GGA GGA ATG TAT AAA 334 Thr Val Ser Ser Val Leu Arg Met Ser Leu Leu Gly Gly Met Tyr Lys

100 105 110

GCC TTT TCA ACA TGT GGA TCT CAT TTG TCG GTT GTC TCT GTT TTA TGG 382 Ala Phe Ser Thr Cys Gly Ser His Leu Ser Val Val Ser Val Leu Trp

115 120 125

CAC AGG TTT TGG GGT ACA CAT AAG CTC TCC ACT TAC TGA CTC TCC AAG 430 His Arg Phe Trp Gly Thr His Lys Leu Ser Thr Tyr * Leu Ser Lys

130 135 140

GAA GAC TGT AGT GGC TTC AGT GAT GTA CAC TGT GGT TAC TCA GAT GCT G 479 Glu Asp Cys Ser Gly Phe Ser Asp Val His Cys Gly Tyr Ser Asp Ala

145 150 155

(2) INFORMATION FOR SEQ ID NO: 34:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 159 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 34:

Ile Cys His Pro Leu Lys Tyr Thr Val Ile Met Asn His Tyr Phe Cys

1 5 10 15

Val Met Leu Leu Leu Phe Ser Val Phe Val Ser Ile Ala His Ala Leu

20 25 30

Phe His Ile Leu Met Val Leu Ile Leu Thr Phe Ser Thr Lys Thr Glu

35 40 45

Ile Pro His Phe Phe Cys Glu Leu Ala His Ile Ile Lys Leu Thr Cys

50 55 60 Ser Asp Asn Phe Ile Asn Tyr Leu Leu Ile Tyr Thr Glu Ser Val Leu

65 70 75 80

Phe Phe Gly Val His Ile Val Gly Ile Ile Leu Ser Tyr lle Tyr Thr

85 90 95

Val Ser Ser Val Leu Arg Met Ser Leu Leu Gly Gly Met Tyr Lys Ala

100 105 no

Phe Ser Thr Cys Gly Ser His Leu Ser Val Val Ser Val Leu Trp His

115 120 125

Arg Phe Trp Gly Thr His Lye Leu Ser Thr Tyr * Leu Ser Lys Glu

130 135 140

Asp Cys Ser Gly Phe Ser Asp Val His Cys Gly Tyr Ser Asp Ala

145 150 155

(2) INFORMATION FOR SEQ ID NO: 35:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 481 base paire

(B) TYPE: nucleic acid

(C) STRANDEDNESS: single

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: CDNA

(iii) HYPOTHETICAL: YES

(iv) ANTI-SENSE: NO

(vi) ORIGINAL SOURCE:

(A) ORGANISM: rat olfactory epithelium

(B) STRAIN: Srpague-Dawley rat

(F) TISSUE TYPE: olfactory epithelium

(vii) IMMEDIATE SOURCE:

(B) CLONE: J20

(ix) FEATURE:

(A) NAME/KEY: CDS

(B) LOCATION: 2..481

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:35:

A ATC TGC TAC CCA CTG AGC TAC CTT CTC ATC ATG ACC TCG GTC GTC 46 Ile Cys Tyr Pro Leu Arg Tyr Leu Leu Ile Met Ser Trp Val Val

1 5 10 15

TGC ACA GCA CTG TCC GTG GCA ATC TGG GTC ATA CGC TTT TGT GCC TCC 94 Cys Thr Ala Leu Ser Val Ala Ile Trp Val Ile Gly Phe Cys Ala Ser

20 25 30

GTT ATA CCT CTC TGC TTC ACG ATC CTC CCA CTC TGT GGT CCT TAC GTC 142 Val Ile Pro Leu Cys Phe Thr Ile Leu Pro Leu Cys Gly Pro Tyr Val

35 40 45

GTT GAT TAT CTT TTC TGC GAG CTG CCC ATC CTT CTG CAC CTG TTC TCC 190 Val Asp Tyr Leu Phe Cys Glu Leu Pro Ile Leu Leu His Leu Phe Cys

50 55 60

ACA GAT ACA TCT CTG CTG GAG NNN NNN NNN NNN NNN NNN NNN NNN NNN 238 Thr Asp Thr Ser Leu Leu Glu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

65 70 75 NNN NNN NNN NNN NNN CCC TTC CTC CTG ATT GTT CTC TCC TAC CTT CGC 286 Xaa Xaa Xaa Xaa Xaa Pro Phe Leu Leu Ile Val Leu Ser Tyr Leu Arg

80 85 90 95

ATC CTG GTG GCT GTG ATA AGA ATA GAC TCA GCT GAG GGC AGA AAA AAG 334 Ile Leu Val Ala Val Ile Arg Ile Asp Ser Ala Glu Gly Arg Lys Lys

100 105 110

GCC TTT TCA ACT TCT GCT TCA CAC TTG GCT GTG GTG ACC ATC TAC TAT 382 Ala Phe Ser Thr Cys Ala Ser His Leu Ala Val Val Thr Ile Tyr Tyr

115 120 125

GGA ACA GGG CTG ATC AGG TAC TTG AGG CCC AAG TCC CTT TAT TCC GCT 430 Gly Thr Gly Leu Ile Arg Tyr Leu Arg Pro Lye Ser Leu Tyr Ser Ala

130 135 140

GAG GGA GAC AGA CTG ATC TCT GTG TTC TAT GCA GTC ATT GGC CCT GCA 478 Glu Gly Asp Arg Leu Ile Ser Val Phe Tyr Ala Val Ile Gly Pro Ala

145 150 155

CTG 481

Leu

160

(2) INFORMATION FOR SEQ ID NO: 36:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 160 amino acids

(B) TYPE: amino acid

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: protein

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 36:

Ile Cye Tyr Pro Leu Arg Tyr Leu Leu Ile Met Ser Trp Val Val Cys

1 5 10 15

Thr Ala Leu Ser Val Ala Ile Trp Val Ile Gly Phe Cys Ala Ser Val

20 25 30

Ile Pro Leu Cys Phe Thr Ile Leu Pro Leu Cys Gly Pro Tyr Val Val

35 40 45

Asp Tyr Leu Phe Cys Glu Leu Pro Ile Leu Leu His Leu Phe Cys Thr

50 55 60

Aep Thr Ser Leu Leu Glu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

65 70 75 80

Xaa Xaa Xaa Xaa Pro Phe Leu Leu Ile Val Leu Ser Tyr Leu Arg Ile

85 90 95

Leu Val Ala Val Ile Arg Ile Asp Ser Ala Glu Gly Arg Lye Lye Ala

100 105 110

Phe Ser Thr Cys Ala Ser His Leu Ala Val Val Thr Ile Tyr Tyr Gly

115 120 125

Thr Gly Leu Ile Arg Tyr Leu Arg Pro Lye Ser Leu Tyr Ser Ala Glu

130 135 140

Gly Aep Arg Leu Ile Ser Val Phe Tyr Ala Val Ile Gly Pro Ala Leu

145 150 155 160

Claims

What is claimed is:

1. An isolated nucleic acid molecule encoding an odorant receptor.

2. An isolated DNA of claim 1.

3. An isolated cDNA of claim 2.

4. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 9.

5. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 10.

6. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 11.

7. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 12.

8. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 13.

9. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 14.

10. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 15.

11. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 16.

12. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 17.

13. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 18.

14. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 19.

15. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 20.

16. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 21.

17. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 22.

18. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 23.

19. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 24.

20. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 25.

21. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 26.

22. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 27.

23. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 28.

24. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 29.

25. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 30.

26. An isolated cDNA of claim 3 wherein the sequence of the cDNA has the nucleotide sequence shown in Figure 31.

27. An isolated cDNA of claim 3 encoding an insect odorant receptor.

28. An isolated cDNA of claim 3 encoding a vertebrate odorant receptor.

29. An isolated cDNA of claim 3 encoding a fish odorant receptor.

30. An isolated cDNA of claim 3 encoding a mammalian odorant receptor.

31. An isolated cDNA of claim 30 wherein the mammalian odorant receptor is a human odorant receptor.

32. An isolated cDNA of claim 30 wherein the mammalian odorant receptor is a rat, dog or mouse odorant receptor.

33. An expression vector comprising the cDNA of claim 3 and the sequence elements necessary for replication and expression in a suitable host.

34. An expression vector comprising the cDNA of any of claims 4-19 and the sequence elements necessary for replication and expression in a suitable host.

35. A purified protein encoding an odorant receptor.

36. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 9.

37. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 10.

38. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 11.

39. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 12.

40. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 13.

41. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 14.

42. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 15.

43. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 16.

44. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 17.

45. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 18.

46. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 19.

47. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 20.

48. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 21.

49. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 22.

50. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 23.

51. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 24.

52. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 25.

53. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 26.

54. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 27.

55. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 28.

56. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 29.

57. A purified protein of claim 35 wherein the amino acid sequence is the sequence in Figure 31.

58. A purified protein of claim 35 encoding an insect odorant receptor.

59. A purified protein of claim 35 encoding a vertebrate odorant receptor.

60. A purified protein of claim 35 encoding a fish odorant receptor.

61. A purified protein of claim 35 encoding a mammalian odorant receptor.

62. A purified protein of claim 61 wherein the mammalian odorant receptor is a human odorant receptor.

63. A purified protein of claim 61 wherein the mammalian odorant receptor is a rat, dog or mouse odorant receptor.

64. A purified protein of claim 35 which has 7 transmembrane regions and whose third cytoplasmic loop from the N-terminus is approximately 17 amino acid long.

65. A method of transforming cells which comprises transfecting a host cell with a suitable expression vector of claim 33.

66. A method of transforming cells which comprises transfecting a host cell with a suitable expression vector of claim 34.

67. Cells transformed by the method of claim 65.

68. Transformed cells of claim 67 wherein the cells are olfactory cells.

69. Transformed cells of claim 67 wherein the cells are non-olfactory cells.

70. A method of identifying a desired odorant ligand comprising contacting transformed non-olfactory cells of claim 69, expressing a known odorant receptor with a series of odorant ligands and determining which ligands bind to the receptors present on the non- olfactory cells.

71. A method of identifying a desired odorant receptor comprising contacting a series of transformed non- olfactory cells of claim 69 with a known odorant ligand and determining which odorant receptor binds with the odorant ligand.

72. A method of detecting an odor which comprises: a) identifying a odorant receptor which binds the desired odorant ligand by the method of claim 71 and; b) imbedding the receptor in a membrane such that when the odorant ligand binds with the receptor identified in a) above, a detectable signal is produced.

73. A method of claim 72 wherein the desired odorant is a pheromone.

74. A method of claim 72 wherein the desired odorant ligand is the vapors emanating from cocaine, marijuana, heroin, hashish, or angel dust.

75. A method of claim 72 wherein the desired odorant ligand is the vapors emanating from gasoline, natural gas or alcohol.

76. A method of claim 72 wherein the desired odorant ligand is the vapors emanating from decayed human flesh.

77. A method of claim 72 wherein the desired odorant ligand is the vapors emanating from explosives, plastic explosives, firearms, or gun powder.

78. A method of claim 72 wherein the desired odorant ligand is toxic fumes, noxious fumes or dangerous fumes.

79. A method of claim 72 wherein the membrane is a cell membrane.

80. A method of claim 72 wherein the membrane is an olfactory cell membrane.

81. A method of claim 72 wherein the membrane is a synthetic membrane.

82. A method of claim 72 wherein the detectable signal is a color change, phosporescence, or radioactivity.

83. A method of quantifying the amount of an odorant ligand present in a sample which comprises the method of claim 72 wherein the detectable signal is quantified.

84. A method of developing fragrances which comprises identifying a desired odorant receptor by the method of claim 71 then contacting non-olfactory cells, which have been transfected with an expression vector containing the cDNA of the desired odorant receptor such that the receptor is expressed upon the surface of the non-olfactory cell, with a series of compounds to determine which ones bind with the receptor.

85. A method of identifying an odorant fingerprint which comprises contacting a series of cells, which have been transformed such that each express a known odorant receptor, with a desired sample and determining the type and quantity of the odorant ligands present in the sample.

86. A method of identifying odorant ligands which inhibit the activity of a desired odorant receptor which comprises contacting the desired odorant receptor with a series of compounds and determining which compounds inhibit the odorant ligand - odorant receptor- interaction.

87. A method of identifying appetite suppressant compounds which comprises identifying odorant ligands by the method of claim 86 wherein the desired odorant receptor is that which is associated with the perception of food.

88. A method of controlling appetite in a subject which comprises contacting the olfactory epithelium of the subject with the odorant ligands identified by the method of claim 87.

89. A nasal spray, to control appetite comprising the compounds identified by the method of claim 87 in a suitable carrier.

90. A method of trapping odors which comprises contacting a membrane which contains multiples of the desired odorant receptor, with a sample such that the desired odorant ligand is absorbed by the binding of the odorant ligand to the odorant receptor.

91. An odor trap employing the method of claim 90.

92. A method of controlling pest populations which comprises identifying odorant ligands by the method of claim 70 which are alarm odorant ligands and spraying the desired area with the identified odorant ligands.

93. A method of controlling a pest population which comprises identifying odorant ligands by the method of claim 70 which interfere with the interaction between the odorant ligands and the odorant receptors which are associated with fertility.

94. A method of claim 92 or 93 wherein the pest population is a population of insects.

95. A method of claim 92 or 93 wherein the pest population is a population of rodents.

96. A method of claim 95 wherein the population of rodents is a population of mice or rats.

97. A method of promoting fertility which comprises employing the method of claim 70 to identify odorant ligands which interact with the odorant receptors associated with fertility and administering the identified odorant ligands to a subject.

98. A method of inhibiting fertility which comprises employing the method of claim 70 to identifying odorant ligands which inhibit the interaction between the odorant ligands and the odorant receptors associated with fertility and administering the identified odorant ligands to a subject.