WO1998037755A1 - Gene de regulation de la reponse de plantes aux pathogenes - Google Patents

Gene de regulation de la reponse de plantes aux pathogenes Download PDF

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Publication number
WO1998037755A1
WO1998037755A1 PCT/US1998/004077 US9804077W WO9837755A1 WO 1998037755 A1 WO1998037755 A1 WO 1998037755A1 US 9804077 W US9804077 W US 9804077W WO 9837755 A1 WO9837755 A1 WO 9837755A1
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WIPO (PCT)
Prior art keywords
seq
dna sequence
isolated dna
lsdl
sequence
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PCT/US1998/004077
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English (en)
Inventor
Jeffery L. Dangl
Robert A. Dietrich
Michael H. Richberg
Petra M. Epple
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The University Of North Carolina At Chapel Hill
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Application filed by The University Of North Carolina At Chapel Hill filed Critical The University Of North Carolina At Chapel Hill
Priority to AU66793/98A priority Critical patent/AU6679398A/en
Priority to EP98908868A priority patent/EP0971579A1/fr
Publication of WO1998037755A1 publication Critical patent/WO1998037755A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
    • C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
    • C12N15/8261—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield
    • C12N15/8271—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance
    • C12N15/8279—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance for biotic stress resistance, pathogen resistance, disease resistance

Definitions

  • Isdl expresses wild type timing of R gene driven HR (Dietrich et al, 1994) ⁇ it is the subsequent spread of cell death which distinguishes the mutant.
  • cell autonomous signals required for R gene function are intact in an Isdl null, but the response to cell non-autonomous signals emanating from cells undergoing HR is perturbed.
  • LSDl functions to limit both the initiation of defense responses and the subsequent extent of the HR.
  • the fact that an Isdl null is hyper-responsive to signals initiating the defense response and HR-like cell death additionally suggests that these pathways are functionally intact in the wild type cell, but require a threshold level of signal for full activation.
  • pSGCGF was made by restricting pGPTV-Hyg with
  • Hindlll and Sad and replacing this fragment with a Hindlll-Sacl fragment containing the polylinker from pIC20H (GenBank accession L08912; provided by Steve Goff, Novartis,
  • the EST 82D11 cDNA sequence was isolated as a Sall-Xbal fragment from pZLl (Newman et al., 1994) and cloned into Xhol-Xbal digested pHyg35S.
  • the sequences of the LSDl cDNA (SEQ ID NOS: 14 and 15) and the 4.5 kb LSDl Xhol-Pstl genomic fragment (SEQ ID NO:13; the longest 5'LSDl cDNA starts at base 1892 of this sequence) are deposited in GenBank as accessions U 87833 and U 87834, respectively. Endpoints of the various LSDl cDNAs isolated are shown in Table3A and examples are provided by SEQ ID NO: 14 (short form from cDNA MG7 as shown in Table 3) and SEQ ID NO: 15 (long form, from cDNA MG8). The polypeptides deduced from these are shown in Fig. 11-12, respectively.
  • Table 3B shows the sizes of each intron deduced from comparison of the sequence shown in SEQ ID NO: 13.
  • Numbers in parentheses refer to the number of isolates of the same clone.
  • Nucleotide numbers at the 5' and 3' ends refer to nucleotide positions from SEQ ID NO: 13.
  • An A at the 3' endpoint can be either an A in the genomic sequence or the first A of the polyA tail. The endpoint marked with an * had no polyA tail.
  • Intron splice junction positions are located at bses 198-199, 260-261, 447-448, 552-553, 692-693, 764-765, and 836-837 in SEQ ID NO: 13.
  • FIG. 1A We constructed a physical contig of these YACs, shown in figure 1A.
  • YAC ends CIC1H1L, yUP5F7R and EG20B4L to isolate genomic phage clones, subcloned fragments form each of these, end-sequenced the subclones, derived primer sequences and developed new CAPS markers (see Tables 1 and 2).
  • the CAPS markers 1H1L-1.6 and 5F7R-1.5 mapped closest to Isdl (1 and 3 recombinants, respectively from 2054 meioses); see Tables 1 and 2 for new CAPS markers).
  • BAC clone 1G5 should contain the gene.
  • transgenic plants were treated with droplets of 2,6-dichloroisonicotinic acid (IN A); 0.3 mg/ml wettable powder containing 25% active ingredient, Uknes et al., 1993a) a potent inducer of SAR and the Isdl phenotype (Dietrich et al., 1994). If the mutation were complemented, then INA treatment should not lead to spreading cell death.
  • Table 4 shows that transgenic plants carrying either the 7kb Xhol fragment or the 4.5 kb Pstl-Xhol ( Figure 3C) all survived this treatment, and are thus complemented for the Isdl mutation.
  • Selfed progeny from a complemented F individual (homozygous Ws-0 alleles through the Isdl interval) were screened by PCR at F for presence of the hygromycin resistance gene and then INA tested, c F parents were identified as hygromycin resistant and heterozygous through the Isdl interval, then selfed and re-screened as hygromycin resistant and homozygous Ws-0 through the Isdl interval at F before INA testing.
  • SEQ ID NO: 13 eight independent cDNAs (Example VII) and completed the sequence of the full 82D11T7 EST sequence.
  • cDNAs we identified two classes expressing open reading frames of either 184 or 189 amino acids (SEQ ID NO: 16 and 17).
  • An alternate splice which adds 61bp to the 5' region of some cDNAs also provides an alternate translation start, hence, the extra five amino acids in SEQ ID NO: 17.
  • the sequences of both cDNA classes matched exactly the genomic sequence except at the positions of 7 introns (see Table 3). Nucleotide 1 of the longest cDNA is at position 1892 in the 4.5 kb Pstl-Xhol genomic sequence (SEQ ID NO: 13 ).
  • the Isdl phenotype can be observed in all cell types examined after initiation of lesion formation (Dietrich et al., 1994).
  • RNA blot analysis of seedlings, stems, leaves and flowers demonstrated that the LSDl gene is expressed constitutively in each of these Arabidopsis tissues (data not shown).
  • the requirement for LSDl activity in these tissues is consistent with the gene's expression pattern.
  • Example XI The LSDl mRNA encodes a novel zinc-finger domain
  • the plant members of this sub-family described to date include the CO gene, which controls transition to flowering (Putterill et al., 1995), a set of related DNA binding proteins (Yanagisawa, 1995; De Paolis et al., 1996) and a gene whose transcription is salt stress- induced (Lippuner et al., 1996). None of these proteins shares with LSDl the consensus homology within the Zn-fingers.
  • the second homology domain is derived from the carboxy 1 portion of LSDl, from residues 129 to 180 ( Figure 6-SEQ ID NO:4). This region of LSDl exhibits homology to three broad classes of regulatory proteins. First, all mammalian insulin receptor substrates; second, a set of animal transcription factors; and third, a maize transcription initiator binding protein.
  • the conceptual LSDl translation product also identified two additional Arabidopsis ESTs via their predicted amino acid homology. Importantly, each has at least one C-x-x-C Zn-fmger and most of the associated consensus residues found in the LSDl internal homologies. They are ESTs 172A7T7 (GenBank R6552)(SEQ ID NO: 58 and 132J21T7 (GenBank T45809). Thus, it is probable that LSDl is the first member of a widely distributed Zn-fmger sub-family in plants, defined by the internal homology within each zinc-finger. The other amino acids in the consensus section are not known to be found in any other zinc finger proteins.
  • Example XII Identification of expressed target sequence tags (EST) and cDNAs containing LSDl -type zinc finger domains was used to search the GenBank database (NCBI). Two Arabidopsis thaliana ESTs (EST132J21T7 and EST 172A7T7) were identified, each of which contains at least two zinc finger domains and most of the associated consensus residues found in the LSDl internal homologies (Dietrich, 1997). These ESTs were ordered from Ohio State University Arabidopsis Biological Resource Stock Center and resequenced. Sequences were analyzed with the Genetics Computer Group programs (Devereaux et al.,1994).
  • RNA isolated from uninduced and P. syringae DC3000 induced Arabidopsis thaliana Col-0 leaf tissue was reverse transcribed.
  • the resulting cDNA population was subcloned unidirectionally into the EcoRI/Xhol - sites of a lambda-Zap II vector using the cDNA-synthesis Kit (Stratagene, La Jolla, CA) according to the
  • the deduced protein (SEQ ID NO:55) consisting of two LSDl -type zinc fmger domains extending from bases 130-195 and 244- 309 of SEQ ID NO:54 (SEQ ID NOS:56-57, respectively).
  • Comparison to EST172A7T7 shows that the EST (SEQ ID NO:58) contains a 124 bp insertion (bases 386-509 after the second zinc finger of SEQ ID NO:58), leading to a different C-terminal.
  • Comparison of these two partial cDNA sequences with the genomic LOLl sequence (see below) demonstrates that they are alternate splice forms from the same gene encoding two related proteins.
  • the erytroid-specific transcription factor EryFl a new finger protein. Cell 58, 877-885.
  • Cytokinesis in the Arabidopsis embryo involves the syntaxin-related KNOLLE gene product. Cell 84, 61-11.
  • the CONSTANS gene of Arabidopsis promotes flowering and encodes a protein showing similarities to zinc finger transcription factors. Cell 80, 847-857.

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  • Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Biophysics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Zoology (AREA)
  • Biochemistry (AREA)
  • Wood Science & Technology (AREA)
  • General Health & Medical Sciences (AREA)
  • General Engineering & Computer Science (AREA)
  • Biotechnology (AREA)
  • Biomedical Technology (AREA)
  • Botany (AREA)
  • Physics & Mathematics (AREA)
  • Cell Biology (AREA)
  • Plant Pathology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Microbiology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Medicinal Chemistry (AREA)
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Abstract

La présente invention concerne des molécules d'ADN codant pour une famille de domaines à doigts de zinc de fixation sur ADN, qui paraît fonctionner de façon à contrôler des niveaux d'un signal dépendant d'un superoxyde et réguler négativement un mécanisme de mort cellulaire de plante comprenant le gène LSD1 de type sauvage, et les protéines LOL1 et LOL2 qui interagissent physiquement avec LSD1, indiquant une fonction qui avec LSD1 régule la réponse des cellules des plantes aux pathogènes.
PCT/US1998/004077 1997-02-28 1998-02-27 Gene de regulation de la reponse de plantes aux pathogenes WO1998037755A1 (fr)

Priority Applications (2)

Application Number Priority Date Filing Date Title
AU66793/98A AU6679398A (en) 1997-02-28 1998-02-27 Plant pathogen response gene
EP98908868A EP0971579A1 (fr) 1997-02-28 1998-02-27 Gene de regulation de la reponse de plantes aux pathogenes

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US3906397P 1997-02-28 1997-02-28
US60/039,063 1997-02-28

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Cited By (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0864650A2 (fr) * 1997-03-11 1998-09-16 Director General of National Institute of Agrobiological Resources, Ministery of Agriculture, Forestry and Fisheries Plantes résistantes au stress
FR2796653A1 (fr) * 1999-07-23 2001-01-26 Agronomique Inst Nat Rech Acides nucleiques codant pour la proteine svn1, application a la modulation des mecanismes de mort cellulaire programmee, notamment la reponse hypersensible (hr), chez les plantes
EP1230345A1 (fr) * 1999-11-17 2002-08-14 Mendel Biotechnology, Inc. Genes resistant au stress environnemental
US7858848B2 (en) 1999-11-17 2010-12-28 Mendel Biotechnology Inc. Transcription factors for increasing yield
US7868229B2 (en) 1999-03-23 2011-01-11 Mendel Biotechnology, Inc. Early flowering in genetically modified plants
CN101130777B (zh) * 2007-01-24 2011-03-23 中国科学院微生物研究所 与水稻生长发育和抗病性相关的编码锌指蛋白的水稻新基因
US7939715B2 (en) 2000-11-16 2011-05-10 Mendel Biotechnology, Inc. Plants with improved yield and stress tolerance
US8106253B2 (en) * 2006-11-15 2012-01-31 Agrigenetics, Inc. Generation of plants with altered protein, fiber, or oil content
US8426678B2 (en) 2002-09-18 2013-04-23 Mendel Biotechnology, Inc. Polynucleotides and polypeptides in plants
US8809630B2 (en) 1998-09-22 2014-08-19 Mendel Biotechnology, Inc. Polynucleotides and polypeptides in plants

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
DIETRICH R. A., ET AL.: "A NOVEL ZINC FINGER PROTEIN IS ENCODED BY THE ARABIDOPSIS LSD1 GENE AND FUNCTIONS AS A NEGATIVE REGULATOR OF PLANT CELL DEATH.", CELL, CELL PRESS, US, vol. 88., 7 March 1997 (1997-03-07), US, pages 685 - 694., XP002910516, ISSN: 0092-8674, DOI: 10.1016/S0092-8674(00)81911-X *
LIPPUNER V., ET AL.: "TWO CLASSES OF PLANT CDNA CLONES DIFFERENTIALLY COMPLEMENT YEAST CALCINEURIN MUTANTS AND INCREASE SALT TOLERANCE OF WILD-TYPE YEAST.", JOURNAL OF BIOLOGICAL CHEMISTRY, AMERICAN SOCIETY FOR BIOCHEMISTRY AND MOLECULAR BIOLOGY, US, vol. 271., no. 22., 31 May 1996 (1996-05-31), US, pages 12859 - 12866., XP002910519, ISSN: 0021-9258, DOI: 10.1074/jbc.271.22.12859 *
PUTTERILL J., ET AL.: "THE CONSTANS GENE OF ARABIDOPSIS PROMOTES FLOWERING AND ENCODES A PROTEIN SHOWING SIMILARITIES TO ZINC FINGER TRANSCRIPTION FACTORS.", NUCLEIC ACIDS RESEARCH, INFORMATION RETRIEVAL LTD., GB, vol. 23., no. 17., 11 September 1995 (1995-09-11), GB, pages 847 - 857., XP002910518, ISSN: 0305-1048 *
YANAGISAWA S.: "A NOVEL DNA-BINDING DOMAIN THAT MAY FORM A SINGLE ZINC FINGER MOTIF", NUCLEIC ACIDS RESEARCH, INFORMATION RETRIEVAL LTD., GB, vol. 23., no. 17., 11 September 1995 (1995-09-11), GB, pages 3403 - 3410., XP002910517, ISSN: 0305-1048, DOI: 10.1093/nar/23.17.3403 *

Cited By (16)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0864650A3 (fr) * 1997-03-11 1999-08-18 Director General of National Institute of Agrobiological Resources, Ministry of Agriculture, Forestry and Fisheries Plantes résistantes au stress
US6310272B1 (en) 1997-03-11 2001-10-30 Director General Of National Institute Of Agrobiological Resources, Ministry Of Agriculture, Forestry And Fisheries Stress resistant plant in which cell death suppressing gene is introduced and method for producing the same
EP0864650A2 (fr) * 1997-03-11 1998-09-16 Director General of National Institute of Agrobiological Resources, Ministery of Agriculture, Forestry and Fisheries Plantes résistantes au stress
US8809630B2 (en) 1998-09-22 2014-08-19 Mendel Biotechnology, Inc. Polynucleotides and polypeptides in plants
US7868229B2 (en) 1999-03-23 2011-01-11 Mendel Biotechnology, Inc. Early flowering in genetically modified plants
FR2796653A1 (fr) * 1999-07-23 2001-01-26 Agronomique Inst Nat Rech Acides nucleiques codant pour la proteine svn1, application a la modulation des mecanismes de mort cellulaire programmee, notamment la reponse hypersensible (hr), chez les plantes
EP1230345A1 (fr) * 1999-11-17 2002-08-14 Mendel Biotechnology, Inc. Genes resistant au stress environnemental
US7858848B2 (en) 1999-11-17 2010-12-28 Mendel Biotechnology Inc. Transcription factors for increasing yield
EP1230345A4 (fr) * 1999-11-17 2005-04-06 Genes resistant au stress environnemental
US9175051B2 (en) 1999-11-17 2015-11-03 Mendel Biotechnology, Inc. Transcription factors for increasing yield
US7939715B2 (en) 2000-11-16 2011-05-10 Mendel Biotechnology, Inc. Plants with improved yield and stress tolerance
US8426678B2 (en) 2002-09-18 2013-04-23 Mendel Biotechnology, Inc. Polynucleotides and polypeptides in plants
US8106253B2 (en) * 2006-11-15 2012-01-31 Agrigenetics, Inc. Generation of plants with altered protein, fiber, or oil content
US8912395B2 (en) 2006-11-15 2014-12-16 Agrigenetics, Inc. Generation of plants with altered protein, fiber, or oil content
US9624501B2 (en) 2006-11-15 2017-04-18 Agrigenetics, Inc. Generation of plants with altered protein, fiber, or oil content
CN101130777B (zh) * 2007-01-24 2011-03-23 中国科学院微生物研究所 与水稻生长发育和抗病性相关的编码锌指蛋白的水稻新基因

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AU6679398A (en) 1998-09-18
EP0971579A1 (fr) 2000-01-19

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