KR20220101913A - 피부 섬유아세포에서 발현하는 노화 관련 후각 수용체 유전자 및 이들의 용도 - Google Patents
피부 섬유아세포에서 발현하는 노화 관련 후각 수용체 유전자 및 이들의 용도 Download PDFInfo
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Abstract
본 발명은 피부 노화의 진단용 조성물에 관한 것으로, 본 발명의 조성물은 다양한 생리적 및 외적 요인으로 인한 현재의 피부 노화 정도를 평가할 뿐 아니라, 향후 이러한 피부 노화가 진행될 유전적 위험성을 높은 신뢰도로 예측함으로써 피부 노화에 대한 개인별 맞춤 치료 전략을 조기에 수립하는 데에 유용하게 이용될 수 있다. 본 발명은 또한 피부 노화 개선제의 스크리닝 방법에 관한 것으로, 본 발명의 스크리닝 방법은 피부 진피(dermis)의 노화에 따라 감소된 진피 섬유아세포의 콜라겐 생성을 복원하여 피부 조직의 결합력 저하, 탄력 감소, 수분 상실 및 장벽기능 손상을 가역적으로 회복시킴으로써 수분 공급과 같은 대증적 요법을 넘어서는 근원적인 피부 노화 억제제를 정확하고 신속하게 탐색할 수 있다.
Description
본 발명은 피부 진피 노화의 지표가 되는 후각 수용체 유전자 및 이를 이용한 피부 노화 진단 방법에 관한 것이다.
피부는 크게 표피(epidermis), 진피(dermis), 피하지방(hypodermis)의 세층으로 구성되어 있다. 표피, 특히 가장 외층을 구성하는 각질층은 피부장벽의 역할을 함으로서 피부로부터 수분과 전해질 손실되는 것을 억제하는 한편, 진피층은 주로 결합조직으로 구성되어 피부의 탄력을 유지하고 구조를 지지하는 역할을 한다. 피부 결합조직에는 세포외기질 단백질인 콜라겐이 가장 많은 양으로 존재하며, 그 밖에 엘라스틴, 피브로넥틴, 인테그린, 피브릴린, 프로테로글리칸 등의 단백질들로 구성되어 있고, 그 사이로 히알루론산이 채워져 있어 피부보습을 담당한다(Fore, J., Ostomy/wound management, 52(9): 24,2006; Yamauchi et al., Journal of Biological Chemistry, 262(24):11428, 1987).
콜라겐은 피부섬유아세포(Dermal fibroblast)에서 새로이 합성된 프로콜라겐으로부터 만들어지는데, 이 프로콜라겐은 효소 반응을 거쳐 피부세포의 세포외 공간으로 분비된 후 삼중나선구조(triple helix configuration)의 마이크로피브릴(microfibril)을 형성하고, 이들은 다시 류신-리치 스몰 프로테오글리칸(leucine-rich small proteoglycans)과 결합하여 피브릴(fibril)을 형성한다. 결과적으로 이렇게 만들어진 피브릴들이 모여 피부의 결합력과 탄력성을 제공하는 콜라겐 섬유를 형성함으로써 피부의 기계적 견고성, 조직의 결합력 및 탄력성 등을 제공한다. 보습을 담당하는 히알루론산 또한 피부섬유아세포에서 히알루론산 합성효소에 의해 만들어지는데, 자신의 무게보다 약 1000배 정도의 물을 끌어당길 수 있는 능력이 있어 보습에 관여한다고 알려져 있다(Veis, A et al., Journal of Biological Chemistry, 264(7):3884, 1989; Hulmes, D. J., Journal of structural biology, 137(1):2, 2002).
피부 노화가 진행됨에 따라 피부 결합조직 중 대부분을 차지하는 단백질인 콜라겐 함량이 감소하는데, 콜라겐은 피부의 장력과 강도를 부여하기 때문에 피부노화에 따른 콜라겐의 감소는 주름생성, 탄력감소와 같은 증상을 일으키는 직접적인 원인으로 지목되고 있다. 피부노화는 크게 생리학적 노화에 의한 내인성 노화, 그리고 환경적인 인자에 의해 일어나는 외인적 노화로 구분된다. 특히 자외선, 염증, 정신적 스트레스, 이 세가지는 콜라겐 양을 현저하게 감소시켜 피부탄력을 감소시키고 주름생성을 촉진시키는 주요한 외인적 노화의 원인으로 손꼽히고 있다 (Bae et al., Experimental dermatology, 19(8):182, 2010; Advani et al., Bone, 20(1):41, 1997; Romana-Souza et al., Mechanisms of ageing and development, 152:63, 2015).
최근 유전정보를 기반으로 한 개인별 맞춤의료가 보건산업의 새로운 패러다임으로 등장하면서 진단검사와 소비자 직접의뢰(DTC; Direct To Consumer) 검사를 수행하는 기업이 점차 증가하고, 유전자 검사 시장이 2017년 58.2억 달러(약 7.7조원)에서 연평균 10.6%씩 성장하여 2024년에는 117.9억 달러(약 14.3조원) 규모로 성장할 것이라는 전망이 보고되었다(생명공학정책연구센터, 2019). 현재 국내의 DTC 유전자 검사 진단 키트는 테라젠바이오와 아모레퍼시픽의‘아이오페 랩 지노 인덱스’, 마크로젠의‘마이지놈스토리 더플러스’, 바이오니아의‘진투라이프’등 다양한 바이오 업체에서 제작되어 대부분 생체 DNA 분석을 통해 타고난 개인의 유전자 발현 상태를 측정하여 혈당, 혈압, 탈모, 콜레스테롤 등 해당 질병에 대해 얼마나 취약한지를 파악하는 것에 초점이 맞춰져 있으나, 선천적 유전 정보가 아닌 현재의 피부 노화 진행 상태를 반영하는 진단 키트는 전무한 실정이다. 또한 시중의 진단 키트는 여러가지 항목에 대한 유전자를 종합 검사하기 위하여 검체를 피부가 아닌 구강상피세포에서 채취하고 있으므로 피부에서 직접 피부노화 상태를 검사할 수 있는 보완 제품의 필요성이 절실한 실정이다.
한편, 후각 수용체(OR; olfactory receptor)는 후각 상피세포에서 주로 발현되는 것으로 알려져 있다. 공기 중의 냄새분자가 후각상피세포의 세포막에 존재하는 후각 수용체와 결합하면, Golf(olfactory G protein)을 우선적으로 활성화시키고 곧이어 AC3(olfactory-related adenylate cyclase)를 활성화시킨다. 후자는 후각세포 내에서 ATP로부터 cAMP(cyclic AMP)의 생성을 촉진시키고, 칼슘이온 채널을 활성화시켜서 궁극적으로 뇌로 신호를 전달하여 냄새 분자를 인식하도록 하는 작용을 한다(Fleischer et al., Frontiers in Cellular Neuroscience 3:9, 2009). 인간에서는 약 350종의 후각 수용체 아형(isoform)이 발현하는 것으로 보고되었고, 이는 전체 지놈 유전자의 1% 이상에 달하는 큰 유전자군으로서, GPCR 중에서도 가장 거대한 군에 해당되는 유전자로 알려져 있다. 단일 후각수용체가 담당하는 냄새 분자는 약 2-3가지이며, 약 100여 가지 후각 수용체의 조합에 따라 사람은 1만 가지 정도의 냄새(또는 냄새 분자)를 구별할 수 있다(Pluznick et al., Proceedings of the National Academy of Sciences, 106(6):2059-2064, 2009).
최근 연구에 의하면 후각 수용체는 후각 조직 뿐 아니라 피부, 폐, 방광, 대장 등 비후각 조직에서 광범위하게 발현되어 신체의 상태에 따라 발현이 다르게 나타나는 것으로 알려져 있다. 예를 들면 아토피 환자의 피부에서 정상인의 피부보다 OR10G7 유전자의 발현이 더 높게 나타났으며(Tham et al., Journal of Allergy and Clinical Immunology, 143(5):1838-1848. 2019), 천식 환자의 폐조직에서 OR2AG2 유전자의 발현이 정상인의 폐조직보다 낮은 것으로 보고되어 현재 질환의 상태 및 민감성을 예측할 수 있는 마커로 제시되었다(Chakraborty et al., Scientific reports 9(1):1-10, 2019). 또한 정상인 방광에 비하여 방광암 환자의 방광에서 OR10H1 유전자의 발현이 현저하게 높아, 방광암의 조기 진단용 바이오마커로 제시된 바 있으며(Weber, L et al., Frontiers in physiology, 9:456,2018), 대장에서 OR7C1 유전자의 발현이 대장암 환자에서 정상인보다 높게 나타나 대장암 진단을 위한 새로운 마커로 제시되었다(Morita et al., Clinical Cancer Research, 22(13):3298-3309, 2016). 따라서, 후각수용체의 이소성 발현은 단순히 후각을 인지하는 것 이상으로 생리학 및 병리학적 상태를 진단 및 대변할 수 있는 마커로서 역할을 할 수 있다는 사실이 최근 2~3년 사이에 보고되고 있으나, 자외선, 염증 및 스트레스과 같은 피부노화 상황에서 진피의 피부섬유아세포의 후각수용체의 발현 변화 여부에 대해서는 아직 밝혀진 바가 없다.
본 명세서 전체에 걸쳐 다수의 논문 및 특허문헌이 참조되고 그 인용이 표시되어 있다. 인용된 논문 및 특허문헌의 개시 내용은 그 전체로서 본 명세서에 참조로 삽입되어 본 발명이 속하는 기술 분야의 수준 및 본 발명의 내용이 보다 명확하게 설명된다.
비특허문헌 1. Romain Duroux et al., A Rose Extract Protects the Skin against Stress Mediators: A Potential Role of Olfactory Receptors. Molecules 25:4743(2020)
본 발명자들은 연령의 증가 또는 다양한 생물학적·비생물학적 스트레스 자극으로 인한 피부 노화의 진행 정도를 정확하게 평가하는 분자 진단 방법을 개발하기 위하여 예의 연구 노력하였다. 그 결과, 외적 및 내적 노화를 야기하는 핵심 노화인자를 인위적으로 가할 경우 피부의 진피(dermis) 조직에서 특정 후각수용체(Olfactory receptor) 유전자의 발현이 유의하게 변화되며, 이들의 발현 수준을 측정할 경우 현재의 피부 노화정도 뿐 아니라 향후 피부 조직의 기계적 견고성, 결합력, 탄력 및 장벽기능이 상실될 유전적 위험성을 높은 신뢰도로 예측할 수 있음을 발견함으로써, 본 발명을 완성하게 되었다.
피부 노화의 진단용 조성물 및 이를 이용한 피부 노화 진단 방법을 제공하는 데 있다.
본 발명의 다른 목적은 피부 노화 개선용 조성물의 스크리닝 방법을 제공하는 데 있다.
본 발명의 다른 목적 및 이점은 하기의 발명의 상세한 설명, 청구범위 및 도면에 의해 보다 명확하게 된다.
본 발명의 일 양태에 따르면, 본 발명은 OR1F2P, OR2A9P, OR2A20P, OR2AK2, OR10A4, OR10A5, OR7E12P, OR2A1/2A42, OR2A7, OR2AE1, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량을 측정하는 제제를 유효성분으로 포함하는 피부 노화의 진단용 조성물을 제공한다.
본 발명자들은 연령의 증가 또는 다양한 생물학적·비생물학적 스트레스 자극으로 인한 피부 노화의 진행 정도를 정확하게 평가하는 분자 진단 방법을 개발하기 위하여 예의 연구 노력하였다. 그 결과, 외적 및 내적 노화를 야기하는 핵심 노화인자를 인위적으로 가할 경우 피부의 진피(dermis) 조직에서 특정 후각수용체(olfactory receptor) 유전자의 발현이 유의하게 변화되며, 이들의 발현 수준을 측정할 경우 현재의 피부 노화정도 뿐 아니라 향후 피부 조직의 기계적 견고성, 결합력, 탄력 및 장벽 기능이 상실될 유전적 위험성을 높은 신뢰도로 예측할 수 있음을 발견하였다.
본 명세서에서 용어“진단”은 피부 노화에 대한 개체의 감수성(susceptibility)의 판정, 현재 개체에서 진행된 피부 노화 정도의 판정 및 향후 피부 노화와 관련한 객체의 예후(prognosis)의 판정을 포함한다. 후술하는 실시예에서 보는 바와 같이, 본 발명에서 발굴된 후각수용체 유전자들의 발현은 피부 조직의 견고성 및 탄력성에 대한 표지자인 콜라겐(collagen)의 발현 변화와 밀접하게 관련되어 있으므로, 용어“피부노화 진단”은“피부 장벽기능의 평가”또는“콜라겐 결핍 관련 피부 질환의 진단”과 동일한 의미로 사용된다.
본 명세서에서 용어“콜라겐 결핍 관련 피부 질환”은 피부 진피조직 내 섬유아세포에 의한 콜라겐 생성이 저하되거나, 또는 MMP-1(collagenase)에 의한 콜라겐 분해 속도가 섬유아세포에 의한 콜라겐 합성 속도를 상회함으로써 나타나는 진피 내 콜라겐의 양적 소실을 원인으로 하는 모든 병적 상태를 포괄하는 의미이다. 각질형성세포(keratinocyte)로 밀집된 피부 표피와는 달리, 피부 진피의 대부분은 세포가 없는 기질 단백질로 이루어져 있다. 콜라겐은 피부 건조 중량의 75%를 차지하는 가장 중요한 기질 단백질로서, 콜라겐의 양적 소실은 전체 기질 단백질의 결핍 및 진피 조직의 구조적 붕괴로 이어져 피부 조직의 결합력 저하, 탄력 감소, 수분 상실 및 장벽기능 손상을 야기한다. 이에, 본 발명의 조성물은 진피 조직 내 유일한 세포성분인 섬유아세포의 후각수용체 유전자 발현양상을 통해 콜라겐 생성의 증감을 정확하게 예측함으로써, 콜라겐 결핍으로 인한 피부 손상 여부 및 향후 손상 발생의 유전적 위험성에 대한 중요한 정보를 제공할 수 있다.
본 발명의 구체적인 구현예에 따르면, 본 발명의 조성물로 진단될 수 있는 피부 노화는 스트레스로 인한 피부 노화이다. 본 명세서에서 용어“스트레스”는 생체의 균형을 붕괴시키는 내외적인 자극에 의해 일어나는 유기체 내의 모든 변화를 포괄하는 의미이다. 스트레스의 원인이 되는 자극은 크게 물리적, 심리적, 생리적 자극으로 분류되며, 물리적 자극은 자연계에 존재하는 기온, 자외선, 화학적 자극 등을, 심리적 자극은 정신적인 고통, 분노, 불안, 긴장 등을, 생리적 자극은 세균이나 바이러스, 알레르기 물질 등을 포함한다. 이에, 본 발명의 조성물로 진단할 수 있는“스트레스로 인한 피부 노화”는 물리적, 심리적, 생리적 자극으로 발생하는 스트레스를 직접 또는 간접적인 원인으로 하는 여하한 피부 노화를 모두 포함한다.
본 명세서에서 용어“진단용 조성물”은 대상체의 피부노화 정도를 판단하거나 향후 피부노화 진행 정도를 예측하기 위해 상기 나열된 후각수용체 유전자 또는 이의 단백질 발현량 측정수단을 포함하는 통합적인 혼합물(mixture) 또는 장비(device)를 의미하며, 이에“진단용 키트”로 표현될 수도 있다.
본 발명의 구체적인 구현예에 따르면, 상기 조성물은 OR1F2P, OR2A9P, OR2A20P, OR2AK2, OR10A4 및 OR10A5로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량을 측정하는 제제를 포함한다. 보다 구체적으로, 상기 나열된 유전자 또는 단백질의 발현량을 측정하는 제제를 포함할 경우 본 발명의 조성물은 자외선 노출에 의한 피부의 노화 정도를 예측한다. 보다 더 구체적으로, 대상체의 시료에서 상기 유전자 중 OR1F2P 또는 이의 인코딩 단백질이 정상 시료보다 고발현되거나, 또는 OR2A9P, OR2A20P, OR2AK2, OR10A4 및 OR10A5로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질이 정상 시료보다 저발현될 경우, 대상체의 피부가 자외선 자극에 의해 노화가 발생한 것으로 판단한다.
본 발명의 구체적인 구현예에 따르면, 상기 자외선 노출에 의한 피부의 노화는 미세주름, 과색소침착, 피부 탄력의 소실 및 수분 결핍으로 구성된 군으로부터 선택되는 하나 이상의 광노화(Photoaging) 현상을 포함하나, 이에 제한되지 않고 자외선 자극을 직접적 또는 간접적인 원인으로 하는 모든 피부 노화 현상을 포괄적으로 포함한다.
본 발명의 구체적인 구현예에 따르면, 상기 조성물은 OR7E12P 유전자 또는 이의 인코딩 단백질의 발현량을 측정하는 제제를 포함한다. 보다 구체적으로, OR7E12P 유전자 또는 이의 인코딩 단백질의 발현량을 측정하는 제제를 포함할 경우 본 발명의 조성물은 조성물은 염증에 의한 피부의 노화 정도를 예측한다. 보다 더 구체적으로, 대상체의 시료에서 OR7E12P 또는 이의 인코딩 단백질이 정상 시료보다 저발현될 경우, 대상체의 피부가 염증 자극에 의해 노화가 발생한 것으로 판단한다.
본 발명의 구체적인 구현예에 따르면, 상기 염증에 의한 피부의 노화는 아토피 피부염, 피부근염, 피부경화증, 천포창, 환상홍반 및 홍반루푸스로 구성된 군으로부터 선택되는 염증성 또는 자가면역성 피부질환으로 인한 피부 노화를 포함하나, 이에 제한되지 않고 과도한 면역반응 또는 염증에 의한 피부 손상을 직접적 또는 간접적인 원인으로 하는 모든 피부 노화 현상을 포괄적으로 포함한다.
본 발명의 구체적인 구현예에 따르면, 상기 조성물은 OR2A1/2A42, OR2A7, OR2AE1, OR2AK2, OR10A4, OR10A5, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량을 측정하는 제제를 포함한다. 보다 구체적으로, 상기 유전자 또는 단백질의 발현량을 측정하는 제제를 포함할 경우 본 발명의 조성물은 피부의 장벽 기능 손상에 의한 피부의 노화 정도를 예측한다. 보다 더 구체적으로, 대상체의 시료에서 상기 유전자 중 하나 이상의 유전자의 또는 이들이 인코딩하는 단백질이 정상 시료보다 저발현될 경우, 대상체의 피부가 장벽 기능 손상에 의해 노화가 발생한 것으로 판단한다.
본 명세서에서 용어“피부 장벽기능 장애(skin barrier dysfunction) 관련 질환”은 콜라겐 소실 및 진피층의 두께 감소로 인해 피부가 외부의 유해분자 투과(permeation)를 차단하고 내부의 수분 유출을 억제하는 장벽기능을 상실함으로써 유발되는 일련의 질환을 포괄하는 의미이다. 장벽기능의 상실은 예를 들어 진피층의 두께 감소로 인한 수분 상실로 인한 피부 노화, 병원균의 침투로 인한 다양한 감염성 질환, 아토피 피부염, 접촉성 피부염과 같은 염증성·자가면역 질환 등의 다양한 병적 상태의 원인이 된다. 본 발명에 따르면, 본 발명자들은 클루코코르티코이드(GC)를 활성화시키는 11β-HSD1(11 beta-hydroxysteroid dehydrogenase type 1)의 발현을 증가시킴으로써 콜라겐 소실과 진피층 두께 감소에 의한 피부 장벽 기능 저하를 야기하는 덱사메타손을 이용하여 피부 스트레스를 유도한 결과, 피부 섬유아세포에서 콜라겐 합성이 감소함과 동시에 상기 나열된 후각수용체 유전자의 발현이 유의하게 감소함을 확인하였다.
본 명세서에서 용어“고발현”은 피부 노화에 대한 양성마커(positive marker)임이 규명된 본 발명의 후각수용체 유전자 또는 이의 인코딩 단백질 발현량이 노화가 진행되지 않은 개체에 비하여 유의하게 높은 경우를 의미하며, 구체적으로는 그 발현량이 정상 개체의 130% 이상인 경우를, 보다 구체적으로는 150% 이상인 경우를, 가장 구체적으로는 170% 이상인 경우를 의미하나, 이를 벗어나는 범위를 제외하는 것은 아니다.
본 명세서에서 용어“저발현”은 피부 노화에 대한 음성마커(positive marker)임이 규명된 본 발명의 후각수용체 유전자 또는 이의 인코딩 단백질 발현량이 노화가 진행되지 않은 개체에 비하여 유의하게 낮은 경우를 의미하며, 구체적으로는 그 발현량이 정상 개체의 약 15% 이상 감소, 보다 구체적으로는 약 30% 이상 감소, 가장 구체적으로는 약 40% 이상 감소한 경우를 의미하나, 이를 벗어나는 범위를 제외하는 것은 아니다.
본 발명의 구체적인 구현예에 따르면, 본 발명의 유전자의 발현량을 측정하는 제제는 상기 유전자의 핵산 분자에 특이적으로 결합하는 프라이머 또는 프로브이다.
본 명세서에서, 용어“핵산 분자”는 DNA(gDNA 및 cDNA) 그리고 RNA 분자를 포괄적으로 포함하는 의미를 갖으며, 핵산 분자에서 기본 구성 단위인 뉴클레오타이드는 자연의 뉴클레오타이드뿐만 아니라, 당 또는 염기 부위가 변형된 유사체 (analogue)도 포함한다(Scheit, Nucleotide Analogs, John Wiley, New York(1980); Uhlman 및 Peyman, Chemical Reviews, 90:543-584(1990)).
본 명세서에서 사용되는 용어“프라이머”는 핵산쇄(주형)에 상보적인 프라이머 연장 산물의 합성이 유도되는 조건, 즉, 뉴클레오타이드와 DNA 중합효소와 같은 중합제의 존재, 적합한 온도와 pH의 조건에서 합성의 개시점으로 작용하는 올리고뉴클레오타이드를 의미한다. 구체적으로는, 프라이머는 디옥시리보뉴클레오타이드 단일쇄이다. 본 발명에서 이용되는 프라이머는 자연(naturally occurring) dNMP(즉, dAMP, dGMP, dCMP 및 dTMP), 변형 뉴클레오타이드 또는 비-자연 뉴클레오타이드를 포함할 수 있으며, 리보뉴클레오타이드도 포함할 수 있다.
본 발명의 프라이머는 타겟 핵산에 어닐링 되어 주형-의존성 핵산 중합효소에 의해 타겟 핵산에 상보적인 서열을 형성하는 연장 프라이머(extension primer)일 수 있으며, 이는 고정화 프로브가 어닐링 되어 있는 위치까지 연장되어 프로브가 어닐링 되어 있는 부위를 차지한다.
본 발명에서 이용되는 연장 프라이머는 타겟 핵산 예를 들어 상기 나열된 후각수용체 유전자의 특정 염기서열에 상보적인 혼성화 뉴클레오타이드 서열을 포함한다. 용어“상보적”은 소정의 어닐링 또는 혼성화 조건하에서 프라이머 또는 프로브가 타겟 핵산 서열에 선택적으로 혼성화할 정도로 충분히 상보적인 것을 의미하며, 실질적으로 상보적(substantially complementary)인 경우 및 완전히 상보적(perfectly complementary)인 경우를 모두 포괄하는 의미이며, 구체적으로는 완전히 상보적인 경우를 의미한다. 본 명세서에서 용어“실질적으로 상보적인 서열”은 완전히 일치되는 서열뿐만 아니라, 특정 서열에 어닐링하여 프라이머 역할을 할 수 있는 범위 내에서, 비교 대상의 서열과 부분적으로 불일치되는 서열도 포함되는 의미이다.
프라이머는, 중합제의 존재 하에서 연장 산물의 합성을 프라이밍시킬 수 있을 정도로 충분히 길어야 한다. 프라이머의 적합한 길이는 다수의 요소, 예컨대, 온도, pH 및 프라이머의 소스(source)에 따라 결정되지만 전형적으로 15-30 뉴클레오타이드이다. 짧은 프라이머 분자는 주형과 충분히 안정된 혼성 복합체를 형성하기 위하여 일반적으로 보다 낮은 온도를 요구한다. 이러한 프라이머의 설계는 타겟 뉴클레오티드 서열을 참조하여 당업자가 용이하게 실시할 수 있으며, 예컨대, 프라이머 디자인용 프로그램(예: PRIMER 3 프로그램)을 이용하여 할 수 있다.
본 명세서에서 용어“프로브”는 특정 뉴클레오타이드 서열에 혼성화될 수 있는 디옥시리보뉴클레오타이드 및 리보뉴클레오타이드를 포함하는 자연 또는 변형되는 모노머 또는 결합을 갖는 선형의 올리고머를 의미한다. 구체적으로, 프로브는 혼성화에서의 최대 효율을 위하여 단일가닥이며, 더욱 구체적으로는 디옥시리보뉴클레오타이드이다. 본 발명에 이용되는 프로브로서, 상기 나열된 각 후각수용체 유전자의 특정 염기서열에 완전하게(perfectly) 상보적인 서열이 이용될 수 있으나, 특이적 혼성화를 방해하지 않는 범위 내에서 실질적으로(substantially) 상보적인 서열이 이용될 수도 있다. 일반적으로, 혼성화에 의해 형성되는 듀플렉스(duplex)의 안정성은 말단의 서열의 일치에 의해 결정되는 경향이 있기 때문에, 타겟 서열의 3’-말단 또는 5’-말단에 상보적인 프로브를 사용하는 것이 바람직하다. 혼성화에 적합한 조건은 Joseph Sambrook, et al., Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory Press, N.Y.(2001) 및 Haymes, B. D., et al., Nucleic Acid Hybridization, A Practical Approach, IRL Press, Washington, D.C.(1985)에 개시된 사항을 참조하여 결정할 수 있다.
본 발명의 구체적인 구현예에 따르면, 본 발명의 단백질의 발현량을 측정하는 제제는 상기 단백질에 특이적으로 결합하는 항체 또는 이의 항원 결합 단편; 또는 상기 단백질에 특이적으로 결합하는 앱타머일 수 있다.
본 발명에 따르면, 본 발명의 후각수용체 단백질을 항원-항체 반응을 이용한 면역분석(immunoassay) 방법에 따라 검출하여 개체의 상피장벽 기능 상실 여부를 분석하는 데 이용될 수 있다. 이러한 면역분석은 종래에 개발된 다양한 면역분석 또는 면역염색 프로토콜에 따라 실시될 수 있다.
예를 들어, 본 발명의 방법이 방사능면역분석 방법에 따라 실시되는 경우, 방사능동위원소(예컨대, C14, I125, P32 및 S35)로 표지된 항체가 이용될 수 있다.
본 명세서에서 용어 "항체"는 포유류의 면역 체계에 의해 생성된, 항원의 에피토프에 결합하는 하나 이상의 가변 도메인을 포함하여 해당 항원을 특이적으로 인식하는 면역글로불린 단백질을 의미한다. 본 발명에서 각각의 후각수용체 단백질을 특이적으로 인식하는 항체는 폴리클로날 또는 모노클로날 항체이며, 바람직하게는 모노클로날 항체이다.
본 발명의 항체는 당업계에서 통상적으로 실시되는 방법들, 예를 들어, 융합 방법(Kohler and Milstein, European Journal of Immunology, 6:511-519 (1976)), 재조합 DNA 방법(미국 특허 제4,816,567호) 또는 파아지 항체 라이브러리 방법(Clackson et al, Nature, 352:624-628(1991) 및 Marks et al, J. Mol. Biol., 222:58, 1-597(1991))에 의해 제조될 수 있다. 항체 제조에 대한 일반적인 과정은 Harlow, E. and Lane, D., Using Antibodies: A Laboratory Manual, Cold Spring Harbor Press, New York, 1999; 및 Zola, H., Monoclonal Antibodies: A Manual of Techniques, CRC Press, Inc., Boca Raton, Florida, 1984에 상세하게 기재되어 있다.
상술한 면역분석 과정에 의한 최종적인 시그널의 강도를 분석함으로써, 상피장벽 기능을 예측할 수 있다. 즉, 개체의 시료에서 양성 마커인 각 후각수용체에 대한 시그널이 정상 시료 보다 강하게 검출되거나 음성 마커인 후각 수용체가 약하게 검출되는 경우, 개체의 피부 장벽기능이 상실되거나 저하되어 피부 노화가 진행된 것으로 판단된다.
본 명세서에서 용어“항원 결합 단편(antigen binding fragment)”은 면역글로불린 전체 구조 중 항원이 결합할 수 있는 폴리펩티드의 일부를 의미하며, 예를 들어 F(ab')2, Fab', Fab, Fv 및 scFv를 포함하나, 이에 제한되는 것은 아니다.
본 명세서에서 용어, "특이적으로 결합(specifically binding)" 은 "특이적으로 인식(specifically recognizing)"과 동일한 의미로서, 항원과 항체(또는 이의 단편)가 면역학적 반응을 통해 특이적으로 상호작용하는 것을 의미한다.
본 발명은 항체 대신 각 후각수용체 단백질에 특이적으로 결합하는 앱타머를 이용할 수도 있다. 본 명세서에서 용어“앱타머”는 특정 표적물질에 높은 친화력과 특이성으로 결합하는 단일 줄기의(single-stranded) 핵산(RNA 또는 DNA) 분자 또는 펩타이드 분자를 의미한다. 앱타머의 일반적인 내용은 Hoppe-Seyler F, Butz K "Peptide aptamers: powerful new tools for molecular medicine". J Mol Med. 78(8):426-30(2000); Cohen BA, Colas P, Brent R . "An artificial cell-cycle inhibitor isolated from a combinatorial library". Proc Natl Acad Sci USA. 95(24):14272-7(1998)에 상세하게 개시되어 있다.
본 발명의 다른 양태에 따르면, 본 발명은 개체로부터 분리된 생물학적 시료 내에서 OR1F2P, OR2A9P, OR2A20P, OR2AK2, OR10A4, OR10A5, OR7E12P, OR2A1/2A42, OR2A7, OR2AE1, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량을 측정하는 단계를 포함하는 피부 노화의 진단에 필요한 정보를 제공하는 방법을 제공한다.
본 발명에서 발굴한 신규 바이오마커인 후각수용체 유전자 및 이를 이용하여 진단될 수 있는 피부 노화에 대해서는 이미 상술하였으므로, 과도한 중복을 피하기 위해 이를 생략한다.
본 발명자들은 상기 나열된 후각수용체 유전자와 피부 노화 간의 상관 관계를 최초로 규명하였다. 이에, 개체 내 상기 후각수용체 유전자 또는 이의 단백질 발현량을 표지자로 하여 개체의 피부 노화 정도를 평가할 수 있다.
본 명세서에서 용어“개체”는 각 후각수용체 유전자 또는 이의 단백질 발현량을 측정하기 위한 시료를 제공하고, 궁극적으로 피부 노화 여부의 분석 대상이 되는 개체를 의미한다. 개체는 제한없이 인간, 마우스, 래트, 기니아 피그, 개, 고양이, 말, 소, 돼지, 원숭이, 침팬지, 비비 또는 붉은털 원숭이를 포함하며, 구체적으로는 인간이다. 본 발명의 조성물은 현재의 피부노화 정도 뿐 아니라 향후 피부노화 과정이 촉진될 유전적 위험성을 예측하기 위한 정보도 제공하므로, 본 발명의“개체”는 피부 노화가 진행되거나 피부 장벽기능이 저하된 환자일 수도 있고 아직 피부 노화가 진행되지 않은 정상개체(healthy subject)일 수도 있다.
본 명세서에서 용어“생물학적 시료”는 인간을 포함한 포유동물로부터 얻어지는, 상술한 후각수용체 유전자를 발현하는 세포를 포함하고 있는 모든 시료로서, 조직, 기관, 세포 또는 세포 배양액을 포함하나, 이에 제한되지 않는다. 구체적으로는, 상기 생물학적 시료는 피부조직 또는 피부조직 유래 세포를 포함하며, 보다 구체적으로는 피부 진피 조직 또는 피부 진피 조직 유래 세포를 포함하고, 가장 구체적으로는 피부 섬유아세포(dermal fibroblast)를 포함한다.
본 발명의 또 다른 양태에 따르면, 본 발명은 다음의 단계를 포함하는 피부 노화 개선용 조성물의 스크리닝 방법을 제공한다:
(a) OR1F2P, OR2A9P, OR2A20P, OR2AK2, OR10A4, OR10A5, OR7E12P, OR2A1/2A42, OR2A7, OR2AE1, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자를 발현하는 세포를 포함하는 생물학적 시료에 후보물질을 접촉시키는 단계;
(b) 상기 시료 내 상기 유전자 또는 상기 유전자가 인코딩하는 단백질의 발현량 또는 활성을 측정하는 단계,
상기 OR1F2P 유전자 또는 이의 인코딩하는 단백질의 발현량 또는 활성이 감소하거나, 상기 OR2A9P, OR2A20P, OR2AK2, OR10A4, OR10A5, OR7E12P, OR2A1/2A42, OR2A7, OR2AE1, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량 또는 활성이 증가한 경우, 상기 후보물질은 피부 노화 개선용 조성물로 판정한다.
본 발명의 구체적인 구현예에 따르면, 본 발명의 스크리닝 방법에 이용되는 세포는 피부 조직 유래 세포이며, 보다 구체적으로는 피부 표피 조직 유래 세포이고, 가장 구체적으로는 피부 섬유아세포(dermal fibroblast)이다.
본 발명의 스크리닝 방법을 언급하면서 사용되는 용어“후보물질”은 상기 나열된 후각수용체를 발현하는 세포를 포함하는 시료에 첨가되어 각 후각수용체 유전자 또는 단백질의 활성 또는 발현량에 영향을 미치는지 여부를 검사하기 위하여 스크리닝에서 이용되는 미지의 물질을 의미한다. 상기 후보물질은 화합물, 뉴클레오타이드, 펩타이드 및 천연 추출물을 포함하나, 이에 제한되는 것은 아니다. 시험물질을 처리한 생물학적 시료에서 각 후각수용체의 발현량 또는 활성을 측정하는 단계는 당업계에 공지된 다양한 발현량 및 활성 측정방법에 의해 수행될 수 있다.
본 명세서에서 용어“활성 또는 발현량의 감소”는 자외선, 염증 자극 또는 스테로이드 제제 자극 등으로 인한 피부 노화 및 장벽기능 손상이 측정 가능한 수준으로 개선될 정도로 피부 노화에 대한 양성마커(positive marker)인 OR1F2P의 생체 내 고유한 기능 또는 발현량이 감소하는 것을 의미한다. 활성(activity)의 감소는 단순한 기능(function)의 감소 뿐 아니라 안정성(stability)의 감소로 기인한 궁극적인 활성 저해를 포함한다. 구체적으로는 후보물질을 처리하지 않은 대조군에 비하여 활성 또는 발현량이 15% 이상 감소한 상태, 보다 구체적으로는 30% 이상 감소한 상태, 가장 구체적으로는 40% 이상 감소한 상태를 의미할 수 있으나, 이를 벗어나는 범위를 제외하는 것은 아니다.
본 명세서에서 용어“활성 또는 발현량의 증가”는 자외선, 염증 자극 또는 스테로이드 제제 자극 등으로 인한 피부 노화 및 장벽기능 손상이 측정 가능한 수준으로 개선될 정도로 피부 노화에 대한 음성마커(positive marker)인 후각수용체의 생체 내 고유한 기능 또는 발현량이 증가하는 것을 의미하며, 구체적으로는 후보물질을 처리하지 않은 대조군에 비하여 활성 또는 발현량이 130%인 경우를, 보다 구체적으로는 150% 이상인 경우를, 가장 구체적으로는 170% 이상인 경우를 의미할 수 있으나, 이를 벗어나는 범위를 제외하는 것은 아니다.
본 발명의 특징 및 이점을 요약하면 다음과 같다:
(a) 본 발명의 조성물은 다양한 생리적 및 외적 요인으로 인한 현재의 피부 노화 정도를 평가할 뿐 아니라, 향후 이러한 피부 노화가 진행될 유전적 위험성을 높은 신뢰도로 예측함으로써 피부 노화에 대한 개인별 맞춤 치료 전략을 조기에 수립하는 데에 유용하게 이용될 수 있다.
(b) 본 발명의 피부 노화 개선용 조성물의 스크리닝 방법은 또한 피부 진피(dermis)의 노화에 따라 감소된 진피 섬유아세포의 콜라겐 생성을 복원하여 피부 조직의 결합력 저하, 탄력 감소, 수분 상실 및 장벽기능 손상을 가역적으로 회복시킴으로써 수분 공급과 같은 대증적 요법을 넘어서는 근원적인 피부 노화 억제제를 정확하고 신속하게 탐색할 수 있다.
도 1은 피부 섬유아세포에서 자외선 처리에 따른 콜라겐 유전자(COL1A1)의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 2는 피부 섬유아세포에서 자외선 처리에 따른 후각수용체 유전자의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 3은 피부 섬유아세포에서 염증 유도 물질인 TNF-α 처리에 따른 COL1A1의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 4는 피부 섬유아세포에서 TNF-α 처리에 따른 후각수용체 유전자의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 5는 피부 섬유아세포에서 스트레스 유도 물질인 덱사메타손(dexamethasone) 처리에 따른 COL1A1의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 6a 및 6b는 피부 섬유아세포에서 덱사메타손 처리에 따른 후각수용체 유전자의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 2는 피부 섬유아세포에서 자외선 처리에 따른 후각수용체 유전자의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 3은 피부 섬유아세포에서 염증 유도 물질인 TNF-α 처리에 따른 COL1A1의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 4는 피부 섬유아세포에서 TNF-α 처리에 따른 후각수용체 유전자의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 5는 피부 섬유아세포에서 스트레스 유도 물질인 덱사메타손(dexamethasone) 처리에 따른 COL1A1의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
도 6a 및 6b는 피부 섬유아세포에서 덱사메타손 처리에 따른 후각수용체 유전자의 발현 변화를 나타낸 그래프이다. 각 값들은 평균 ± 표준오차(SEM)로 나타내었으며, 그룹 간 유의한 차이가 있는 경우 별표(*p < 0.05)로 표시하였다.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하고자 한다. 이들 실시예는 오로지 본 발명을 보다 구체적으로 설명하기 위한 것으로, 본 발명의 요지에 따라 본 발명의 범위가 이들 실시예에 의해 제한되지 않는다는 것은 당업계에서 통상의 지식을 가진 자에 있어서 자명할 것이다.
실시예
실시예 1: 자외선 자극에 의한 피부노화 관련 유전자 및 후각수용체 유전자의 발현 변화
실험방법
1) 피부 섬유아세포 배양
실험에 사용한 피부 섬유아세포(dermal fibroblast cell line, Hs68)는 ATCC사(Manassas, VA, USA)에서 구입하여 사용하였으며, 10% 우태혈청(fetal bovine serum; HyClone, Logan, UT, USA)과 1% 항생제(페니실린 및 스트렙토마이신; Gibco)를 첨가한 DMEM(Dulbecco’s modified eagle medium; Hyclone) 배지를 사용하여 37℃, 5% CO₂조건 하에서 배양하였다.
2) 자외선 조사
피부 섬유아세포에 자외선을 조사하기 위해 UV 조사기(Model CL-1000M; UVP, Upland, CA, USA)를 이용하였다. 세포의 배양액을 제거한 후 PBS(phosphate buffered saline; WelGENE, Daegu, Korea)로 세척하고 세포를 살짝 덮을 만큼의 PBS를 넣은 상태에서 자외선(20 mJ/cm2)을 조사하였다.
3) RNA 추출 및 실시간 PCR
피부 섬유아세포에서 자외선 조사에 의해 변화하는 피부노화 관련 유전자와 후각수용체 유전자 발현 패턴을 정량적으로 분석하기 위해 실시간 PCR을 이용하였다. 세포에서 RNA를 추출하기 위해 배양액을 제거하고 차가운 PBS로 세척한 다음 TRIzol 시약(Invitrogen, Carlsbad, CA, USA)을 사용하여 총 RNA를 분리한 후 분광광도계(NanoQuant Infinite M200 Pro; Tecan, MSwitzerland)로 RNA를 정량하였다. 추출한 RNA로부터 cDNA를 합성하기 위해 역전사효소(SuperScript Ⅳ reverse transcriptase; Invitrogen, Carlsbad, CA, USA)를 사용하였다.
실시간 PCR은 사이버그린 믹스(SYBR green supermix; BioRad, Hercules, CA, USA)에 프라이머와 cDNA를 넣고 CFX Connect™ 실시간 PCR Detection System (BioRad)을 이용하여 95℃에서 10초, 58℃에서 15초, 60℃에서 15초의 과정을 36회 반복하여 수행하였으며 결과는 Bio-Rad CFX Manager 3.0으로 확인하였다. 각 유전자의 발현은 GAPDH(Glyceraldehyde 3-phosphate dehydrogenase)의 발현에 대해 표준화하여 비교·분석하였다. 검출하고자 하는 유전자 특이적인 프라이머를 제작하여 사용하였으며, 사용된 프라이머의 염기서열은 표 1에 나타내었다.
유전자 | 프라이머 염기서열 (5′→ 3′) | |
COL1A1 | F | ACATGTTCAGCTTTGTGGACC |
R | TGTACGCAGGTGATTGGTGG | |
OR1F2P | F | CTGAGGGTCCCATCCACAAAG |
R | GGGAGGATGAAGTTCTGAAATATGG | |
OR2A9P | F | CATCCTGAGGAGCAGCAGAAG |
R | AGGCAATAATTGGGCATCAAGAG | |
OR2A20P | F | TCCCTGTTCTATGTCTTCACCC |
R | CAGGAGGCATTCAGTATGTGC | |
OR2AK2 | F | TCTATGCAACCACTCTCTTTACC |
R | AAATACTGCCACCGCCTTATCC | |
OR10A4 | F | CCCTTGCACTACCCAGTCATC |
R | TCAGAGCCTCCAGTTCAAACAC | |
OR10A5 | F | CAAAGGACACGGGCCAAAC |
R | CAGCAATGCGAGTATAGGAACAC | |
GAPDH | F | AAATCAAGTGGGGCGATGC |
R | AGGGGGCAGAGATGATGACC |
4) 통계 분석
모든 실험결과는 평균 ± 표준오차로 표시하였고, SPSS Statistics (version 24.0, Chicago, IL, USA)를 사용하여 독립표본 t-검정(student's t-test)을 실시하여 p < 0.05 수준에서 유의성을 검증하였다.
실험결과
1) 자외선 조사에 의한 피부노화 관련 유전자의 감소
자외선을 조사한 피부 섬유아세포에서 콜라겐 유전자(COL1A1)의 발현이 자외선을 조사하지 않은 피부 섬유아세포에 비해 유의적으로 감소하였다(도 1).
2) 자외선 조사에 의한 후각수용체 유전자의 발현 변화
자외선을 처리한 피부 섬유아세포에서 후각수용체 유전자 OR1F2P의 발현이 자외선을 처리하지 않은 피부 섬유아세포에 비해 유의적으로 증가하였다(도 2a). 한편, 후각수용체 유전자 OR2A9P, OR2A20P, OR2AK2, OR10A4 및 OR10A5의 발현은 유의적으로 감소하였다(도 2b).
실시예 2: 염증 자극에 의한 피부노화 관련 유전자 및 후각수용체 유전자의 발현 변화
실험방법
1) 피부 섬유아세포 배양
실험에 사용한 피부 섬유아세포(human dermal fibroblast cell line, Hs68)는 실시예 1에서와 동일한 방법으로 배양하였다.
2) 염증유도물질 처리
피부 섬유아세포에서 염증상황을 유발하기 위해 TNF-α(tumor necrosis factor-α) 10 ng/ml를 처리하여 12시간 동안 배양하였다. 음성 대조군은 TNF-α의 용매(DMSO; dimethyl sulfoxide; Sigma-Aldrich, St. Louis, MO, USA)만 처리 후 12시간 동안 배양하였다.
3) RNA 추출 및 실시간 PCR
피부 섬유아세포에서 염증유도물질 처리에 의해 변화하는 피부노화 관련 유전자와 후각수용체 유전자 발현패턴을 정량적으로 분석하기 위해 실시간 PCR을 이용하였다. RNA 추출 및 실시간 PCR은 실시예 1에서와 동일한 방법으로 진행되었다. 검출하고자 하는 유전자 특이적인 프라이머를 제작하여 사용하였으며, 실시예 2에서 사용된 프라이머의 염기서열은 표 2에 나타내었다.
유전자 | 프라이머 염기서열 (5′→ 3′) | |
OR7E12P | F | TTGCTGAGTGTGGATTGCTCT |
R | TGATGAGTAGGTTCCCCAGC |
4) 통계 분석
모든 실험결과는 평균 ± 표준오차로 표시하였고, SPSS Statistics (version 24.0, Chicago, IL, USA)를 사용하여 독립표본 t 검정(student's t test)을 실시하여 p < 0.05 수준에서 유의성을 검증하였다.
실험결과
1) 염증 자극에 의한 피부노화 관련 유전자의 감소
염증유도물질(TNF-α)을 처리한 피부 섬유아세포에서 콜라겐 유전자(COL1A1)의 발현이 염증유도물질을 처리하지 않은 피부 섬유아세포에 비해 유의적으로 감소하였다(도 3).
2) 염증 자극에 의한 후각수용체 유전자의 발현 변화
염증유도물질(TNF-α)을 처리한 피부 섬유아세포에서 후각수용체 유전자 OR7E12P의 발현이 염증유도물질을 처리하지 않은 피부 섬유아세포에 비해 유의적으로 감소하였다(도 4).
실시예 3: 스트레스 자극에 의한 피부노화 관련 유전자 및 후각수용체 유전자의 발현 변화
실험방법
1) 피부 섬유아세포 배양
실험에 사용한 피부 섬유아세포(human dermal fibroblast cell line, Hs68)는 실시예 1에서와 동일한 방법으로 배양하였다.
2) 스트레스 유도물질 처리
피부 섬유아세포에서 스트레스 상황을 유발하기 위해 합성 코티졸(cortisol)인 덱사메타손(dexamethasone)을 1 μM의 농도로 처리하여 12시간 동안 배양하였다. 음성 대조군은 덱사메타존의 용매인 DMSO만 처리 후 12시간 동안 배양하였다.
3) RNA 추출 및 실시간 PCR
피부 섬유아세포에서 덱사메타손 처리에 의해 변화하는 피부노화 관련 유전자와 후각수용체 유전자 발현패턴을 정량적으로 분석하기 위해 실시간 PCR을 이용하였다. RNA 추출 및 실시간 PCR은 실시예 1에서와 동일한 방법으로 진행되었다. 검출하고자 하는 유전자 특이적인 프라이머를 제작하여 사용하였으며, 실시예 3에서 사용된 프라이머의 염기서열은 표 3에 나타내었다.
유전자 | 프라이머 염기서열 (5′→ 3′) | |
OR2A1/2A42 | F | GCATGACGCAGACCTTTCTCT |
R | GCAGTCTTAGGATGAGAACCA | |
OR2A7 | F | GACCTCATTCAACCTCACAATC |
R | ATTTCATTCAGCGGCAAACAC | |
OR2AE1 | F | TGTGGCAGAAGAATCAGACC |
R | GAGAATGGTGAGGGTGTTGC | |
OR51B4 | F | CTGCTAGACCAGAGGGAGATTG |
R | TGTTCATCACTCGGGAATTGGTAAG | |
OR51B5 | F | ACTGTGGTAATAACAGAAACAGAAAG |
R | ACCAGTCAATAGGAAGGGATGG | |
OR51I1 | F | CCATGCTCAACCCTATCATCTAC |
R | GGCCTGGGATTTATGGAAGAAC | |
OR52D1 | F | TCTCCCCCTTCATCTTCTTGCT |
R | GAGCCACAGTTAGCCCATAGA |
4) 통계 분석
모든 실험결과는 평균 ± 표준오차로 표시하였고, SPSS Statistics (version 24.0, Chicago, IL, USA)를 사용하여 독립표본 t 검정(student's t test)을 실시하여 p < 0.05 수준에서 유의성을 검증하였다.
실험결과
1) 스트레스 자극에 의한 피부노화 관련 유전자의 발현 변화
스트레스 유도물질(덱사메타손)을 처리한 피부 섬유아세포에서 COL1A1 유전자의 발현이 스트레스 유도물질을 처리하지 않은 피부 섬유아세포에 비해 유의적으로 감소하였다(도 5).
2) 스트레스 자극에 의한 후각수용체 유전자의 발현 변화
스트레스 유도물질(덱사메타손)을 처리한 피부 섬유아세포에서 후각수용체 유전자 OR2A1/2A42, OR2A7, OR2AE1, OR2AK2, OR10A4, OR10A5, OR51B4, OR51B5, OR51I1 및 OR52D1의 발현이 스트레스 유도물질을 처리하지 않은 피부 섬유아세포에 비해 유의적으로 감소하였다(도 6a 및 도 6b).
후각수용체 | 서열번호 및 Genbank 접근번호 | |
아미노산 서열 | 뉴클레오타이드 서열 | |
OR1F2P | 비코딩 유전자 | 제1서열 (NR_002169.1) |
OR2A1/2A42 | 제2서열 (NP_001005287.1) | 제3서열 (NM_030908.2) |
OR2A7 | 제4서열 (NP_001005328.1) | 제5서열 (NM_001005328.2) |
OR2A9P | 비코딩 유전자 | 제6서열 (NR_002157.1) |
OR2A20P | 비코딩 유전자 | 제7서열 (NR_002158.1) |
OR2AE1 | 제8서열 (NR_002169.1) | 제9서열 (NM_001005276.1) |
OR2AK2 | 제10서열 (NR_002169.1) | 제11서열 (NM_001004491.1) |
OR7E12P | 비코딩 유전자 | 제12서열 (NR_044999.1) |
OR10A4 | 제13서열 (NP_997069.2) | 제14서열 (NM_207186.2) |
OR10A5 | 제15서열 (NP_835462.1) | 제16서열 (NM_178168.1) |
OR51B4 | 제17서열 (NP_149419.2) | 제18서열 (NM_001171454.1) |
OR51B5 | 제19서열 (NP_001005567.2) | 제20서열 (NM_001005567.3) |
OR51I1 | 제21서열 (NP_001005169.1) | 제22서열 (NM_001005169.1) |
OR52D1 | 제23서열 (NP_001005163.1) | 제24서열 (NM_001005163.2) |
이상으로 본 발명의 특정한 부분을 상세히 기술하였는 바, 당업계의 통상의 지식을 가진 자에게 있어서 이러한 구체적인 기술은 단지 바람직한 구현예일 뿐이며, 이에 본 발명의 범위가 제한되는 것이 아닌 점은 명백하다. 따라서, 본 발명의 실질적인 범위는 첨부된 청구항과 그의 등가물에 의하여 정의된다고 할 것이다.
<110> Industry-Academic Cooperation Foundation, Yonsei University
<120> Skin Aging-Related Olfactory Receptor Genes Expressed in Dermal
Fibroblasts and Use Thereof
<130> HPC-9687
<160> 56
<170> KoPatentIn 3.0
<210> 1
<211> 985
<212> DNA
<213> Homo sapiens
<400> 1
atgtgtatgt ttctgaattc actgtcttct atgcagctgg gtccagacat atgagaggga 60
caaaccagtg agtgtctccg agttcctcct cttgggactc tccaggcagc cccagcagca 120
gcatctcctc tttgtgttct tcctcagcat gtacctggcc actgtcctgg ggaacctgct 180
catcatcctg gccataagca tagactcccg cctgcacacc cccatgtact tcttcctcag 240
caacatgtcc tttgtggaca actgcttctc caccaccgtc cccaagatgc tggccaatca 300
catactcagg actcaaacca tctccttctc tggctgtctc atgcagatgt attttatcag 360
tgagcttgct gacatggaca atttcctcct ggctgtgatg gcctatgacc gctttgtcgc 420
cgtgtgccgc cccttacatt acacagcaaa gatgacccat cagctctgtg ccctgctggt 480
cactggatca tgggtggttg ccaactcgaa tgctctgctg cacaccctgc tgatggctcg 540
actctcattc tgtgcagaca acaccatccc ccacatcttc tgcgatgtga ctcccctcct 600
gaaactctcc tgttcagaca cacacctcag tgaagtgatg attcttactg aggctgccct 660
agtcacgatc accccatttc tttgcctcct ggcttcctat atgcacatca cctgcgttgt 720
cctgagggtc ccatccacaa agggaagatg gaaagccttc tccacctgtg gctcccacct 780
ggctgtggtt ctcctcttct atggcaccat catgtctcca tatttcagaa cttcatcctc 840
ccactcagct cagagagata tagcagctgc tgtgaggttc acagtggtga ctcccgtgat 900
gaatcctttg atctacagcc tgaggaacaa ggacataaaa ggggctcttg taaaagtggt 960
tgctgtgaaa tttttttctg ttcaa 985
<210> 2
<211> 310
<212> PRT
<213> Homo sapiens
<400> 2
Met Gly Glu Asn Gln Thr Met Val Thr Glu Phe Leu Leu Leu Gly Phe
1 5 10 15
Leu Leu Gly Pro Arg Ile Gln Met Leu Leu Phe Gly Leu Phe Ser Leu
20 25 30
Phe Tyr Ile Phe Thr Leu Leu Gly Asn Gly Ala Ile Leu Gly Leu Ile
35 40 45
Ser Leu Asp Ser Arg Leu His Thr Pro Met Tyr Phe Phe Leu Ser His
50 55 60
Leu Ala Val Val Asp Ile Ala Tyr Thr Arg Asn Thr Val Pro Gln Met
65 70 75 80
Leu Ala Asn Leu Leu His Pro Ala Lys Pro Ile Ser Phe Ala Gly Cys
85 90 95
Met Thr Gln Thr Phe Leu Cys Leu Ser Phe Gly His Ser Glu Cys Leu
100 105 110
Leu Leu Val Leu Met Ser Tyr Asp Arg Tyr Val Ala Ile Cys His Pro
115 120 125
Leu Arg Tyr Ser Val Ile Met Thr Trp Arg Val Cys Ile Thr Leu Ala
130 135 140
Val Thr Ser Trp Thr Cys Gly Ser Leu Leu Ala Leu Ala His Val Val
145 150 155 160
Leu Ile Leu Arg Leu Pro Phe Ser Gly Pro His Glu Ile Asn His Phe
165 170 175
Phe Cys Glu Ile Leu Ser Val Leu Arg Leu Ala Cys Ala Asp Thr Trp
180 185 190
Leu Asn Gln Val Val Ile Phe Ala Ala Cys Val Phe Phe Leu Val Gly
195 200 205
Pro Pro Ser Leu Val Leu Val Ser Tyr Ser His Ile Leu Ala Ala Ile
210 215 220
Leu Arg Ile Gln Ser Gly Glu Gly Arg Arg Lys Ala Phe Ser Thr Cys
225 230 235 240
Ser Ser His Leu Cys Val Val Gly Leu Phe Phe Gly Ser Ala Ile Ile
245 250 255
Met Tyr Met Ala Pro Lys Ser Arg His Pro Glu Glu Gln Gln Lys Val
260 265 270
Phe Phe Leu Phe Tyr Ser Phe Phe Asn Pro Thr Leu Asn Pro Leu Ile
275 280 285
Tyr Ser Leu Arg Asn Gly Glu Val Lys Gly Ala Leu Arg Arg Ala Leu
290 295 300
Gly Lys Glu Ser His Ser
305 310
<210> 3
<211> 4746
<212> DNA
<213> Homo sapiens
<400> 3
ctcttctgag atccttgagt ccaaatgtgg gagctaacca ggctgcctct ttttacttta 60
ccatgaattc ctcctagaca tgttgtagct cctggtcttg gttaactttc acatccacta 120
caaacattgg gaaatggggg aaaatcagac aatggtcaca gagttcctcc tactgggatt 180
tctcctgggc ccaaggattc agatgctcct ctttgggctc ttctccctgt tctatatctt 240
caccctgctg gggaacgggg ccatcctggg gctcatctca ctggactcca gactccacac 300
ccccatgtac ttcttcctct cacacctggc tgtcgtcgac atcgcctaca cccgcaacac 360
ggtgccccag atgctggcga acctcctgca tccagccaag cccatctcct ttgctggctg 420
catgacgcag acctttctct gtttgagttt tggacacagc gaatgtctcc tgctggtgct 480
gatgtcctac gatcgttacg tggccatctg ccaccctctc cgatactccg tcatcatgac 540
ctggagagtc tgcatcaccc tggccgtcac ttcctggacg tgtggctccc tcctggctct 600
ggcccatgtg gttctcatcc taagactgcc cttctctggg cctcatgaaa tcaaccactt 660
cttctgtgaa atcctgtctg tcctcaggct ggcctgtgct gacacctggc tcaaccaggt 720
ggtcatcttt gcagcctgcg tgttcttcct ggtggggcca cccagcctgg tgcttgtctc 780
ctactcgcac atcctggcgg ccatcctgag gatccagtct ggggagggcc gcagaaaggc 840
cttctccacc tgctcctccc acctctgcgt ggtgggactc ttctttggca gtgccatcat 900
catgtacatg gcccccaagt cccgccatcc tgaggagcag caaaaggtct tttttctatt 960
ttacagtttt ttcaacccaa cacttaaccc cctgatttac agcctgagga acggagaggt 1020
caagggtgcc ctgaggagag cactgggcaa ggaaagtcat tcctaactgg tgtgacattt 1080
gactctccct cctcagtcat ctcctggaat cttggtacca aataccacct aagttcacta 1140
ctctctttat atctgagact gaatgaacca agagactctg caaagcattc ctttttcctg 1200
cctgggaagt atttagtttt tgatgcattt gttatactta acatttttta atttaagcac 1260
ttattgagtt gagaatgtcg gggaaagatt tattttgtac actgctatga gtccagaagt 1320
ttaaaacaga cactccacct gtgacttagt caggtatgct cagtagtaga ggaatagcag 1380
ttattaccag ataaagtcat atctataaaa agtttttaaa aatacagcca caggcagaga 1440
ctccagaacc cattgatacc tctgtccatt ttcatcctca tttggatgtt tcccctcaat 1500
tgtacttcct ttcttataag cagtaaatgt acctaagtgc ctacttcagc ttggagggaa 1560
attgattttt ataaaatcct acaaactgac aatttctgtt gtcagagaga gtgaatccat 1620
aaacattcag ctggcaatct gaagcaatta cacaacaaaa tccactttcc aacctgccct 1680
gagactgtga tggtcacttg tttctactct tctcacctcc tccaggggac tgtctatgat 1740
tgtctctaaa taagcacatg ctcttcactt agaggaagtt agtaccacca cacaactttt 1800
cactgctggg gacgaatgga aagccccaca cagagacaat tcaatggaaa aaagcgtttt 1860
aataataatg ttaatagtta gcattattat aaattactct gcaccatcca ctgcacattg 1920
ataggtactt tatagagata acctcagttt atcattatag ccaattcatt tcccattatt 1980
atcccatttt atagatgtga acttcaagca gatggtaaat gttagactca tacccaaaca 2040
atctggctcc caagccttct ttacttaacc aggacaatgt gggactttat tagcaacatt 2100
tcaaattaag aataagtcta accaatccta acgtaggata tgttaagtga atgtgggttt 2160
cttccccatc tagaatgtgc catgatgagg ccaggagagt cagagattat ttatggaagg 2220
cttcttatcc tgtttgacga gtcctgtggg catggctgca aaggaggtta agagattcag 2280
aggaacacag aacacagaca gggtgggagt agaggctgtt ctcaatgaga ttagagagta 2340
agtttagaac tatacttggt tctgggtggt aagatccaga acaaagacat ctctagttca 2400
catacaacac tcacatgctc acgtacatgc gtgtgcacac acacacacac acacacacat 2460
atacgcacag tgcttagtga agcactattc caagaacctt acaagtaaga gctctttaag 2520
tcctcataaa aaccctatga ggtagagact attactacta tttttaggcg agaaaacaga 2580
actatagaga gattatataa tttaaccaat gctcccagct aataagtacc tagtccagaa 2640
tcaaacttag gcaatatgac tgctgaaccc ataatcgtaa ccattatata aaaatcaaag 2700
agatcatgaa atttccaaag atgaagagga ggagagatga aaagcaattt tcaatctcta 2760
gccaaagata ggcattgggc tccagacaga aaagtactta aagctggggg aaaaacaaac 2820
aaacaaacaa acaaacaaac aaaaaacagc tttttttata ctggtgcgtg aaatgagatg 2880
aggtggtagc aagaaggaac tgtgactcca agggtcagag acggaggagg agcctcttga 2940
cgggcaggat ggtgaaacca aaaacaatgg ataaaacttt taagttgatg gttcatcaat 3000
tgaagaacat tagaaagtgt ttttgtaaat tatctagccg ggcatggcaa cgtgtgccta 3060
tatccccagc tattcgggag gctgaggcaa gaggatcact tgagccaggg attttgaggc 3120
tgcagtgagc catgatcgca ccactgcatt ccagcctggg tgacagagtg agaccttgtg 3180
tccaacaata ataaaaatga aataatttat taaatttttt aaaattaccc aaactggata 3240
tacaatttgt aattccattg ctgccgaaca gtttttaaac ggttgtttag tttcacggca 3300
gatgctgcat actgcattgc aactaacact gggaagggat acattcaagt ggcaggacaa 3360
aggtgaggag ctctaatacc aggggtaggt ttcaagggat gaatactccc ctggtaccag 3420
aggagcagca ggaaatcatg gtgtgcagaa accatgctca ggcggttgcc tgtgtggacc 3480
ctaaattctg tgtgtgcagt cctctcttag gattggccca ggcatccaag aagcccacga 3540
tgctgcctac cccttaaata cacaaggtgc gtaccggagg ccctggttct agttccagat 3600
gtaccatttg ctagctgtgt tttttgcaga gaagtccctt aaactgctta agcttcagtc 3660
cttcatctga aaattatgca tatggaagct gtttaggaca ccacagagtg ccactgaaaa 3720
cccaaagcaa gatgcaaatg gacacatttt ttttctgaag acttaaaagt gctatgtgaa 3780
cataagataa actgttatga ttaccctgct gtcagaagtt attttaatcc tgtcagggtt 3840
gctaaacttt ctgatctgga accaaaattg ctctttaaac aggaagagca ttggtgctgg 3900
taggtcacct acgcctttgc cctcatggct tccccttgtc ccccctagag gcacaggagg 3960
ccactcactc ttggagggca tagctggacc aggagcatga gctgtggccc agactcaggg 4020
aacctctgtg agtcagctgg caaaggctcc cggggagcct gtctctgaat ctggtcctag 4080
cccgttcact tggaaaaaac aggagacttc tcccaaggtc aaggaattca gattttggtt 4140
ttggtttcca actttttgta atcctcaacc tgggatttac agtgcatgtc tcaattatgt 4200
ttcattcatt gcttcacaaa gggaaggaca agcactggtt catctcttta cacctataca 4260
ttcctagcac agagtgagta ctcacaaaaa acatacggaa tacatagagg acactttatc 4320
tgttgacccc atttccaaat tttcagtgtt gagccatctt ctgatttcta aacttcttta 4380
aatccctcta attttaggtt atactacagt agaaatttaa ttgtagctga taattattga 4440
gcacttacta tgtggttggc actgcataaa atgctttaga aaaattctta catcaaatat 4500
gtggagtagg tactaatact atcctaacat taaagggggg gaaataaagt ttagtgtaat 4560
taattaattt attcaaggtc gccaaatcat tcagtggtag aaacatgaat caagaccagg 4620
cagtctgact gtaaataaat tctgagccac tctcttgcag aggacctcct gatattcatc 4680
tcctgaatgg ttggcaacag gcctgttcat agtagacaca caataaactt tgactaaatg 4740
aacaaa 4746
<210> 4
<211> 310
<212> PRT
<213> Homo sapiens
<400> 4
Met Gly Asp Asn Ile Thr Ser Ile Thr Glu Phe Leu Leu Leu Gly Phe
1 5 10 15
Pro Val Gly Pro Arg Ile Gln Met Leu Leu Phe Gly Leu Phe Ser Leu
20 25 30
Phe Tyr Val Phe Thr Leu Leu Gly Asn Gly Thr Ile Leu Gly Leu Ile
35 40 45
Ser Leu Asp Ser Arg Leu His Ala Pro Met Tyr Phe Phe Leu Ser His
50 55 60
Leu Ala Val Val Asp Ile Ala Tyr Ala Cys Asn Thr Val Pro Arg Met
65 70 75 80
Leu Val Asn Leu Leu His Pro Ala Lys Pro Ile Ser Phe Ala Gly Arg
85 90 95
Met Met Gln Thr Phe Leu Phe Ser Thr Phe Ala Val Thr Glu Cys Leu
100 105 110
Leu Leu Val Val Met Ser Tyr Asp Leu Tyr Val Ala Ile Cys His Pro
115 120 125
Leu Arg Tyr Leu Ala Ile Met Thr Trp Arg Val Cys Ile Thr Leu Ala
130 135 140
Val Thr Ser Trp Thr Thr Gly Val Leu Leu Ser Leu Ile His Leu Val
145 150 155 160
Leu Leu Leu Pro Leu Pro Phe Cys Arg Pro Gln Lys Ile Tyr His Phe
165 170 175
Phe Cys Glu Ile Leu Ala Val Leu Lys Leu Ala Cys Ala Asp Thr His
180 185 190
Ile Asn Glu Asn Met Val Leu Ala Gly Ala Ile Ser Gly Leu Val Gly
195 200 205
Pro Leu Ser Thr Ile Val Val Ser Tyr Met Cys Ile Leu Cys Ala Ile
210 215 220
Leu Gln Ile Gln Ser Arg Glu Val Gln Arg Lys Ala Phe Cys Thr Cys
225 230 235 240
Phe Ser His Leu Cys Val Ile Gly Leu Phe Tyr Gly Thr Ala Ile Ile
245 250 255
Met Tyr Val Gly Pro Arg Tyr Gly Asn Pro Lys Glu Gln Lys Lys Tyr
260 265 270
Leu Leu Leu Phe His Ser Leu Phe Asn Pro Met Leu Asn Pro Leu Ile
275 280 285
Cys Ser Leu Arg Asn Ser Glu Val Lys Asn Thr Leu Lys Arg Val Leu
290 295 300
Gly Val Glu Arg Ala Leu
305 310
<210> 5
<211> 2062
<212> DNA
<213> Homo sapiens
<400> 5
atcacgtgtt gtttaatgat gcggatacgt tctgagaaat gtgcggtcaa gtgattctga 60
cattgtgtga acatccagaa aggacttcac ggggatatgg gggacaatat aacatccatc 120
acagagttcc tcctactggg atttcccgtt ggcccaagga ttcagatgct cctctttggg 180
ctcttctccc tgttctacgt cttcaccctg ctggggaacg ggaccatact ggggctcatc 240
tcactggact ccagactgca cgcccccatg tacttcttcc tctcacacct ggcggtcgtc 300
gacatcgcct acgcctgcaa cacggtgccc cggatgctgg tgaacctcct gcatccagcc 360
aagcccatct cctttgcggg ccgcatgatg cagacctttc tgttttccac ttttgctgtc 420
acagaatgtc tcctcctggt ggtgatgtcc tatgatctgt acgtggccat ctgccacccc 480
ctccgatatt tggccatcat gacctggaga gtctgcatca ccctcgcggt gacttcctgg 540
accactggag tccttttatc cttgattcat cttgtgttac ttctaccttt acccttctgt 600
aggccccaga aaatttatca ctttttttgt gaaatcttgg ctgttctcaa acttgcctgt 660
gcagataccc acatcaatga gaacatggtc ttggccggag caatttctgg gctggtggga 720
cccttgtcca caattgtagt ttcatatatg tgcatcctct gtgctatcct tcagatccaa 780
tcaagggaag ttcagaggaa agccttctgc acctgcttct cccacctctg tgtgattgga 840
ctcttttatg gcacagccat tatcatgtat gttggaccca gatatgggaa ccccaaggag 900
cagaagaaat atctcctgct gtttcacagc ctctttaatc ccatgctcaa tccccttatc 960
tgtagtctta ggaactcaga agtgaagaat actttgaaga gagtgctggg agtagaaagg 1020
gctttatgaa aaggattatg gcattgtgac tgacagtgac ctaggaagtt acatcattga 1080
gcggttctta acccatctct gcactggtgg gacctctgcc ctcaatggac atgagaatta 1140
tctgagacat ttatttaaaa tggagctatc tcctgcccta cctttaaatg actgatttca 1200
gcagatgtgg gatgaaattc tagaaattga tctcttcaag ttgtgctgca gccactccac 1260
accaggacaa taccccttac aatatcctca ttagcttttg atccagtccc atacctccca 1320
taatgttttc ctcaaaacac tggtcccttc agaggacttt aaaaataagt tctatagtca 1380
aataagtttg agacttactt cacatcagat gcctctgtct agggatcaca attcatatta 1440
gcatagtgaa tgctgtaaat atttctctag gaaagaataa ttctatacca gctttaagcc 1500
agtgttttct aaacttatgt gagcacataa aatttccttt gtaatactta gtaacagttt 1560
cctggaacag gagatcttga tattaattga ctcattgtga atacttccta cagccccctt 1620
ctagggcaga atgatttctt ttttccttgc aaagtgagcc tctaaagaga tgtagttctg 1680
agcattatgc ctcgatcagt ctgtaaaaat ccgggttctg tttggataca gaccgtgagg 1740
gaccctgtct ctactgttca atggtagatc atctaaagaa aataaatgca atcctgtcct 1800
tcatcatgga tcggcattcc tgctatagaa gcttcaggag atgacctcat tcaacctcac 1860
aatcttctaa atttgagatt tttaagaaat atgattcagt atacatctgc ctgtgtttgc 1920
cgctgaatga aatcaatcta atttttctaa ttcagggttt tcaagataaa cccagatttc 1980
tcaagaaaga aaaatctaga aagatttcaa aatccaccac ctacacactt aaaagttgta 2040
gcagcttctt atgccatggg aa 2062
<210> 6
<211> 985
<212> DNA
<213> Homo sapiens
<400> 6
tggactccag actccacacc cccatgtact tcttcctctc acacctggcc gtcgtcaaca 60
tcgcctatgc ctgcaacaca gtgccccaga tgctggtgaa cctcctgcat ccagccaagc 120
ccatctcctt tgctggctgc atgacataga cctttctctt tttgagtttt gcacatactg 180
aatgcctcct gttggtgctg atgtcctacg atcggtacgt ggccatctgc caccctctcc 240
gatatttcat catcatgacc tggaaagtct gcatcactct ggccatcact tcctggacat 300
gtggctccct cctggctatg gtccatgtga gcctcatcct aagactgccc ttttgtgggc 360
ctcgtgaaat caaccacttc ttctgtgaaa tcctgtctgt cctcaggctg gcctgtgctg 420
atacctggct caaccaggtg gtcatctttg cagcctgcat gttcatcctg gtgggaccac 480
tctgcctggt gctggtctcc tactcacaca tcctggcggc catcctgagg atccagtctg 540
gggagggccg cagaaaggcc ttctccacct gctcctccca cctctgcgta gtgggactct 600
tctttggcag cgccatcgtc atgtacatgg cccctaagtc ccgccatcct gaggagcagc 660
agaaggtcct ttttctattt tacagttctt tcaacccgat gctaaacccc ctgatttaca 720
acctgaggaa tgtagaggtc aagggtgccc tgaggagagc actgtgcaag gaaagtcatt 780
cctaagaggt gtgacatttg aactgccagc ctcagttgtc acgtggactc ttgatgccca 840
attattgcct caatccagaa aagtttactt ctctttatct gtgctttact gacagaaggg 900
caagtcttct ctcgtttttt gcagataaaa ttttagatgt gttgcattca ttgggtttct 960
atgagatgtg gttttatcag acaat 985
<210> 7
<211> 930
<212> DNA
<213> Homo sapiens
<400> 7
atggtgaaaa atcagacaat ggtcacagag ttcctcctac tgggatttct cctgggccca 60
aggattcaga tgctcctctt tgggctcttc tccctgttct atgtcttcac cctgctgggg 120
aatgggacca tcctggggct catctcactg gactccagac tccacacccc catgtacttc 180
ttcctctcac acctggccgt cgtcaacatc gcctatgcct gcaacacagt gccccagatg 240
ctggtgaacc tcctgcatcc agccaagccc atctcctttg ctggctgcat gacatagacc 300
tttctctttt tgagttttgc acatactgaa tgcctcctgt tggtgctgat gtcctacgat 360
cggtacgtgg ccatctgcca ccctctccga tatttcatca tcatgacctg gaaagtctgc 420
atcactctgg ccatcacttc ctggacatgt ggctccctcc tggctatggt ccatgtgagc 480
ctcatcctaa gactgccctt ttgtgggcct cgtgaaatca accacttctt ctgtgaaatc 540
ctgtctgtcc tcaggctggc ctgtgctgat acctggctca accaggtggt catctttgca 600
gcctgcatgt tcatcctggt gggaccactc tgcctggtgc tggtctccta ctcacacatc 660
ctggcggcca tcctgaggat ccagtctggg gagggccgca gaaaggcctt ctccacctgc 720
tcctcccacc tctgcgtagt gggactcttc tttggcagcg ccatcgtcat gtacatggcc 780
cctaagtccc gccatcctga ggagcagcag aaggtccttt ttctatttta cagttctttc 840
aacccgatgc taaaccccct gatttacaac ctgaggaatg tagaggtcaa gggtgccctg 900
aggagagcac tgtgcaagga aagtcattcc 930
<210> 8
<211> 323
<212> PRT
<213> Homo sapiens
<400> 8
Met Trp Gln Lys Asn Gln Thr Ser Leu Ala Asp Phe Ile Leu Glu Gly
1 5 10 15
Leu Phe Asp Asp Ser Leu Thr His Leu Phe Leu Phe Ser Leu Thr Met
20 25 30
Val Val Phe Leu Ile Ala Val Ser Gly Asn Thr Leu Thr Ile Leu Leu
35 40 45
Ile Cys Ile Asp Pro Gln Leu His Thr Pro Met Tyr Phe Leu Leu Ser
50 55 60
Gln Leu Ser Leu Met Asp Leu Met His Val Ser Thr Ile Ile Leu Lys
65 70 75 80
Met Ala Thr Asn Tyr Leu Ser Gly Lys Lys Ser Ile Ser Phe Val Gly
85 90 95
Cys Ala Thr Gln His Phe Leu Tyr Leu Cys Leu Gly Gly Ala Glu Cys
100 105 110
Phe Leu Leu Ala Val Met Ser Tyr Asp Arg Tyr Val Ala Ile Cys His
115 120 125
Pro Leu Arg Tyr Ala Val Leu Met Asn Lys Lys Val Gly Leu Met Met
130 135 140
Ala Val Met Ser Trp Leu Gly Ala Ser Val Asn Ser Leu Ile His Met
145 150 155 160
Ala Ile Leu Met His Phe Pro Phe Cys Gly Pro Arg Lys Val Tyr His
165 170 175
Phe Tyr Cys Glu Phe Pro Ala Val Val Lys Leu Val Cys Gly Asp Ile
180 185 190
Thr Val Tyr Glu Thr Thr Val Tyr Ile Ser Ser Ile Leu Leu Leu Leu
195 200 205
Pro Ile Phe Leu Ile Ser Thr Ser Tyr Val Phe Ile Leu Gln Ser Val
210 215 220
Ile Gln Met Arg Ser Ser Gly Ser Lys Arg Asn Ala Phe Ala Thr Cys
225 230 235 240
Gly Ser His Leu Thr Val Val Ser Leu Trp Phe Gly Ala Cys Ile Phe
245 250 255
Ser Tyr Met Arg Pro Arg Ser Gln Cys Thr Leu Leu Gln Asn Lys Val
260 265 270
Gly Ser Val Phe Tyr Ser Ile Ile Thr Pro Thr Leu Asn Ser Leu Ile
275 280 285
Tyr Thr Leu Arg Asn Lys Asp Val Ala Lys Ala Leu Arg Arg Val Leu
290 295 300
Arg Arg Asp Val Ile Thr Gln Cys Ile Gln Arg Leu Gln Leu Trp Leu
305 310 315 320
Pro Arg Val
<210> 9
<211> 972
<212> DNA
<213> Homo sapiens
<400> 9
atgtggcaga agaatcagac ctctctggca gacttcatcc ttgaggggct cttcgatgac 60
tcccttaccc accttttcct tttctccttg accatggtgg tcttccttat tgcggtgagt 120
ggcaacaccc tcaccattct cctcatctgc attgatcccc agcttcatac accaatgtat 180
ttcctgctca gccagctctc cctcatggat ctgatgcatg tctccacaat catcctgaag 240
atggctacca actacctatc tggcaagaaa tctatctcct ttgtgggctg tgcaacccag 300
cacttcctct atttgtgtct aggtggtgct gaatgttttc tcttagctgt catgtcctat 360
gaccgctatg ttgccatctg tcatccactg cgctatgctg tgctcatgaa caagaaggtg 420
ggactgatga tggctgtcat gtcatggttg ggggcatccg tgaactccct aattcacatg 480
gcgatcttga tgcacttccc tttctgtggg cctcggaaag tctaccactt ctactgtgag 540
ttcccagctg ttgtgaagtt ggtatgtggc gacatcactg tgtatgagac cacagtgtac 600
atcagcagca ttctcctcct cctccccatc ttcctgattt ctacatccta tgtcttcatc 660
cttcaaagtg tcattcagat gcgctcatct gggagcaaga gaaatgcctt tgccacttgt 720
ggctcccacc tcacggtggt ttctctttgg tttggtgcct gcatcttctc ctacatgaga 780
cccaggtccc agtgcactct attgcagaac aaagttggtt ctgtgttcta cagcatcatt 840
acgcccacat tgaattctct gatttatact ctccggaata aagatgtagc taaggctctg 900
agaagagtgc tgaggagaga tgttatcacc cagtgcattc aacgactgca attgtggttg 960
ccccgagtgt ag 972
<210> 10
<211> 335
<212> PRT
<213> Homo sapiens
<400> 10
Met Asn Ile Ser Asp Val Ile Ser Phe Asp Ile Leu Val Ser Ala Met
1 5 10 15
Lys Thr Gly Asn Gln Ser Phe Gly Thr Asp Phe Leu Leu Val Gly Leu
20 25 30
Phe Gln Tyr Gly Trp Ile Asn Ser Leu Leu Phe Val Val Ile Ala Thr
35 40 45
Leu Phe Thr Val Ala Leu Thr Gly Asn Ile Met Leu Ile His Leu Ile
50 55 60
Arg Leu Asn Thr Arg Leu His Thr Pro Met Tyr Phe Leu Leu Ser Gln
65 70 75 80
Leu Ser Ile Val Asp Leu Met Tyr Ile Ser Thr Thr Val Pro Lys Met
85 90 95
Ala Val Ser Phe Leu Ser Gln Ser Lys Thr Ile Arg Phe Leu Gly Cys
100 105 110
Glu Ile Gln Thr Tyr Val Phe Leu Ala Leu Gly Gly Thr Glu Ala Leu
115 120 125
Leu Leu Gly Phe Met Ser Tyr Asp Arg Tyr Val Ala Ile Cys His Pro
130 135 140
Leu His Tyr Pro Met Leu Met Ser Lys Lys Ile Cys Cys Leu Met Val
145 150 155 160
Ala Cys Ala Trp Ala Ser Gly Ser Ile Asn Ala Phe Ile His Thr Leu
165 170 175
Tyr Val Phe Gln Leu Pro Phe Cys Arg Ser Arg Leu Ile Asn His Phe
180 185 190
Phe Cys Glu Val Pro Ala Leu Leu Ser Leu Val Cys Gln Asp Thr Ser
195 200 205
Gln Tyr Glu Tyr Thr Val Leu Leu Ser Gly Leu Ile Ile Leu Leu Leu
210 215 220
Pro Phe Leu Ala Ile Leu Ala Ser Tyr Ala Arg Val Leu Ile Val Val
225 230 235 240
Phe Gln Met Ser Ser Gly Lys Gly Gln Ala Lys Ala Val Ser Thr Cys
245 250 255
Ser Ser His Leu Ile Val Ala Ser Leu Phe Tyr Ala Thr Thr Leu Phe
260 265 270
Thr Tyr Thr Arg Pro His Ser Leu Arg Ser Pro Ser Arg Asp Lys Ala
275 280 285
Val Ala Val Phe Tyr Thr Ile Val Thr Pro Leu Leu Asn Pro Phe Ile
290 295 300
Tyr Ser Leu Arg Asn Lys Glu Val Thr Gly Ala Val Arg Arg Leu Leu
305 310 315 320
Gly Tyr Trp Ile Cys Cys Arg Lys Tyr Asp Phe Arg Ser Leu Tyr
325 330 335
<210> 11
<211> 1008
<212> DNA
<213> Homo sapiens
<400> 11
atgaacattt cagatgtcat ctcctttgat attttggttt cagccatgaa aacaggaaat 60
caaagttttg ggacagattt tctacttgtt ggtcttttcc aatatggctg gataaactct 120
cttctctttg tcgtcattgc caccctcttt acagttgctc tgacaggaaa tatcatgctg 180
atccacctca ttcgactgaa caccagactc cacactccaa tgtactttct gctcagtcag 240
ctctccatcg ttgacctcat gtacatctcc accacagtgc ccaagatggc agtcagcttc 300
ctctcacaga gtaagaccat tagatttttg ggctgtgaga ttcaaacgta tgtgttcttg 360
gcccttggtg gaactgaagc ccttctcctt ggttttatgt cttatgatcg ctatgtagct 420
atctgtcacc ctttacatta tcctatgctt atgagcaaga agatctgctg cctcatggtt 480
gcatgtgcat gggccagtgg ttctatcaat gctttcatac atacattgta tgtgtttcag 540
cttccattct gtaggtctcg gctcattaac cactttttct gtgaagttcc agctctacta 600
tcattggtgt gtcaggacac ctcccagtat gagtatacag tcctcctgag tggacttatt 660
atcttgctac taccattcct agccattctg gcttcctatg ctcgtgtgct tattgtggta 720
ttccagatga gctcaggaaa aggacaggca aaagctgttt ccacttgttc ctcccacctg 780
attgtggcaa gcctgttcta tgcaaccact ctctttacct acacaaggcc acactccttg 840
cgttcccctt cacgggataa ggcggtggca gtattttaca ccattgtcac acctctactg 900
aacccattta tctacagcct gagaaataag gaagtgacgg gggcagtgag gagactgttg 960
ggatattgga tatgctgtag aaaatatgac ttcagatctc tgtattga 1008
<210> 12
<211> 1209
<212> DNA
<213> Homo sapiens
<400> 12
tgagcttgtc agaactacaa aatattgctg agtgtggatt gctctgaaac ctgcaaacat 60
tactggtgaa ttgcttctat ccaaaatgca gacacaatgc tgggtattgg tttacttgtt 120
tcggattttt caaccctctt ttccaggcaa aagagggttg tatccaaatg atacagaccc 180
acagagtcta acagatgtct ctatattcct cctcctcaaa ctctcagagg atccagaact 240
gcagcaggtc gtcgctgggc tgttcctgtc catgtgcctg gtcacggtgc tggggaacct 300
actcatcatc ctggccgtca gccctgactc ccacctccac acccccatgt acttgttcct 360
ctccaacctg tccttgcctg acatcggttt cacctccacc acggtcccca agatgattgt 420
ggacatccag tctcacagca gagtcatctc ctatgcaggc tgcctgactc agatgtctct 480
ctttgccatt tttggaggta tggaagagag acatgctcct gagtgtgatg gcctatgacc 540
ggtttgtagc catctgtcac cctctatatt gttcagccat ctttaacccg tgtttctgtg 600
gcttcctaga tttgttgtct tttttttttt ttttctcagt ctttcagact cccagctgca 660
caacttgatt gccttacaaa tgacctgctt caaggatgtg gaaattccta atttcttctg 720
ggaaccttct caactctccc atcttgcatg ttgtgacacc ttcaccagga acatcagtat 780
ttccctgctg ccatatttgg ttttcttccc atcttgggga cccttttctc ttactgtaaa 840
attgtttcct ccattctgag ggtttcatca tcaggtggga agtataaacc ttctccacct 900
gtgggtctca cctgtcagtt gtttgctgat tttatggaac aggcattgga gggtacctcg 960
gttcagatgt gtcatcttcc ccgagaaagg gtgcagtggc ctcagtgatg tacatggtgg 1020
tcacccccat gctgaacccc ttcatctaca gcctgagaaa cagggatatg aaaagtgtcc 1080
tgcggcggcc gcatggcagc acagtctaat ctcaacatct tcttatctgt tccattcctt 1140
ttgtagggtg gtttaaaaaa ggcgccaagg tcaaataaga atgacatcac agggtgaaca 1200
cccactgtg 1209
<210> 13
<211> 315
<212> PRT
<213> Homo sapiens
<400> 13
Met Met Trp Glu Asn Trp Thr Ile Val Ser Glu Phe Val Leu Val Ser
1 5 10 15
Phe Ser Ala Leu Ser Thr Glu Leu Gln Ala Leu Leu Phe Leu Leu Phe
20 25 30
Leu Thr Ile Tyr Leu Val Thr Leu Met Gly Asn Val Leu Ile Ile Leu
35 40 45
Val Thr Ile Ala Asp Ser Ala Leu Gln Ser Pro Met Tyr Phe Phe Leu
50 55 60
Arg Asn Leu Ser Phe Leu Glu Ile Gly Phe Asn Leu Val Ile Val Pro
65 70 75 80
Lys Met Leu Gly Thr Leu Ile Ile Gln Asp Thr Thr Ile Ser Phe Leu
85 90 95
Gly Cys Ala Thr Gln Met Tyr Phe Phe Phe Phe Phe Gly Ala Ala Glu
100 105 110
Cys Cys Leu Leu Ala Thr Met Ala Tyr Asp Arg Tyr Val Ala Ile Cys
115 120 125
Asp Pro Leu His Tyr Pro Val Ile Met Gly His Ile Ser Cys Ala Gln
130 135 140
Leu Ala Ala Ala Ser Trp Phe Ser Gly Phe Ser Val Ala Thr Val Gln
145 150 155 160
Thr Thr Trp Ile Phe Ser Phe Pro Phe Cys Gly Pro Asn Arg Val Asn
165 170 175
His Phe Phe Cys Asp Ser Pro Pro Val Ile Ala Leu Val Cys Ala Asp
180 185 190
Thr Ser Val Phe Glu Leu Glu Ala Leu Thr Ala Thr Val Leu Phe Ile
195 200 205
Leu Phe Pro Phe Leu Leu Ile Leu Gly Ser Tyr Val Arg Ile Leu Ser
210 215 220
Thr Ile Phe Arg Met Pro Ser Ala Glu Gly Lys His Gln Ala Phe Ser
225 230 235 240
Thr Cys Ser Ala His Leu Leu Val Val Ser Leu Phe Tyr Ser Thr Ala
245 250 255
Ile Leu Thr Tyr Phe Arg Pro Gln Ser Ser Ala Ser Ser Glu Ser Lys
260 265 270
Lys Leu Leu Ser Leu Ser Ser Thr Val Val Thr Pro Met Leu Asn Pro
275 280 285
Ile Ile Tyr Ser Ser Arg Asn Lys Glu Val Lys Ala Ala Leu Lys Arg
290 295 300
Leu Ile His Arg Thr Leu Gly Ser Gln Lys Leu
305 310 315
<210> 14
<211> 995
<212> DNA
<213> Homo sapiens
<400> 14
ccagtgaagt cctgggcaga agaatgatgt gggaaaactg gacaattgtc agtgaatttg 60
ttctcgtgag cttctcagcc ctgtccactg agcttcaggc tctactgttt ctccttttct 120
tgaccattta cttggttact ttaatgggca atgtcctcat catcctggtc actatagctg 180
actctgcact acaaagtcct atgtacttct tcctcagaaa cttgtccttc ctggagatag 240
gtttcaactt ggtcattgtg cccaagatgc tggggaccct gatcattcaa gacacaacca 300
tctccttcct tggatgtgcc actcagatgt atttcttctt cttttttggg gctgctgagt 360
gctgcctcct ggccaccatg gcatatgacc gctacgtggc catctgtgac cccttgcact 420
acccagtcat catgggccac atatcctgtg cccagctggc agctgcctct tggttctcag 480
ggttttcagt ggccactgtg caaaccacat ggattttcag tttccctttt tgtggcccca 540
acagggtgaa ccacttcttc tgtgacagcc ctcctgttat tgcactggtc tgtgctgaca 600
cctctgtgtt tgaactggag gctctgacag ccactgtcct attcattctc tttcctttct 660
tgctgatcct gggatcctat gtccgcatcc tctccactat cttcaggatg ccgtcagctg 720
aggggaaaca tcaggcattc tccacctgtt ccgcccacct cttggttgtc tctctcttct 780
atagcactgc catcctcacg tatttccgac cccaatccag tgcctcttct gagagcaaga 840
agctgctgtc actctcttcc acagtggtga ctcccatgtt gaaccccatc atctacagct 900
caaggaataa agaagtgaag gctgcactga agcggcttat ccacaggacc ctgggctctc 960
agaaactatg attggcttag atggaaactg aaggg 995
<210> 15
<211> 317
<212> PRT
<213> Homo sapiens
<400> 15
Met Ala Ile Gly Asn Trp Thr Glu Ile Ser Glu Phe Ile Leu Met Ser
1 5 10 15
Phe Ser Ser Leu Pro Thr Glu Ile Gln Ser Leu Leu Phe Leu Thr Phe
20 25 30
Leu Thr Ile Tyr Leu Val Thr Leu Lys Gly Asn Ser Leu Ile Ile Leu
35 40 45
Val Thr Leu Ala Asp Pro Met Leu His Ser Pro Met Tyr Phe Phe Leu
50 55 60
Arg Asn Leu Ser Phe Leu Glu Ile Gly Phe Asn Leu Val Ile Val Pro
65 70 75 80
Lys Met Leu Gly Thr Leu Leu Ala Gln Asp Thr Thr Ile Ser Phe Leu
85 90 95
Gly Cys Ala Thr Gln Met Tyr Phe Phe Phe Phe Phe Gly Val Ala Glu
100 105 110
Cys Phe Leu Leu Ala Thr Met Ala Tyr Asp Arg Tyr Val Ala Ile Cys
115 120 125
Ser Pro Leu His Tyr Pro Val Ile Met Asn Gln Arg Thr Arg Ala Lys
130 135 140
Leu Ala Ala Ala Ser Trp Phe Pro Gly Phe Pro Val Ala Thr Val Gln
145 150 155 160
Thr Thr Trp Leu Phe Ser Phe Pro Phe Cys Gly Thr Asn Lys Val Asn
165 170 175
His Phe Phe Cys Asp Ser Pro Pro Val Leu Lys Leu Val Cys Ala Asp
180 185 190
Thr Ala Leu Phe Glu Ile Tyr Ala Ile Val Gly Thr Ile Leu Val Val
195 200 205
Met Ile Pro Cys Leu Leu Ile Leu Cys Ser Tyr Thr Arg Ile Ala Ala
210 215 220
Ala Ile Leu Lys Ile Pro Ser Ala Lys Gly Lys His Lys Ala Phe Ser
225 230 235 240
Thr Cys Ser Ser His Leu Leu Val Val Ser Leu Phe Tyr Ile Ser Ser
245 250 255
Ser Leu Thr Tyr Phe Trp Pro Lys Ser Asn Asn Ser Pro Glu Ser Lys
260 265 270
Lys Leu Leu Ser Leu Ser Tyr Thr Val Val Thr Pro Met Leu Asn Pro
275 280 285
Ile Ile Tyr Ser Leu Arg Asn Ser Glu Val Lys Asn Ala Leu Ser Arg
290 295 300
Thr Phe His Lys Val Leu Ala Leu Arg Asn Cys Ile Pro
305 310 315
<210> 16
<211> 954
<212> DNA
<213> Homo sapiens
<400> 16
atggctatag gaaactggac agaaataagt gaatttatcc tcatgagctt ctcttcccta 60
cctactgaaa tacagtcatt gctcttcctg acatttctaa ctatctattt ggttactctg 120
aagggaaaca gcctcatcat tctggttacc ctagctgacc ccatgctaca cagccccatg 180
tacttcttcc tcagaaactt atctttcctg gagattggct tcaacctagt cattgtgccc 240
aaaatgctgg ggaccctgct tgcccaggac acaaccatct ccttccttgg ctgtgccact 300
cagatgtatt tcttcttctt ctttggggta gctgaatgct tcctcctggc taccatggca 360
tatgaccgct atgtggccat ctgcagtccc ttgcactacc cagtcatcat gaaccaaagg 420
acacgggcca aactggctgc tgcttcctgg ttcccaggct ttcctgtagc tactgtgcag 480
accacatggc tcttcagttt tccattctgt ggcaccaaca aggtgaacca cttcttctgt 540
gacagcccgc ctgtgctgaa gctggtctgt gcagacacag cactgtttga gatctacgcc 600
atcgtcggaa ccattctggt ggtcatgatc ccctgcttgc tgatcttgtg ttcctatact 660
cgcattgctg ctgctatcct caagatccca tcagctaaag ggaagcataa agccttctct 720
acgtgctcct cacacctcct tgttgtctct cttttctata tatcttctag cctcacctac 780
ttctggccta aatcaaataa ttctcctgag agcaagaagt tgttatcatt atcctacact 840
gttgtgactc ccatgttgaa ccccattatc tacagcttga gaaatagcga ggtgaagaat 900
gccctcagca ggaccttcca caaggtccta gccctcagaa actgtatccc atag 954
<210> 17
<211> 310
<212> PRT
<213> Homo sapiens
<400> 17
Met Trp Tyr Asn Asn Ser Ala Gly Pro Phe Leu Leu Thr Gly Phe Leu
1 5 10 15
Gly Ser Glu Ala Val His Tyr Arg Ile Ser Met Ser Phe Phe Val Ile
20 25 30
Tyr Phe Ser Val Leu Phe Gly Asn Gly Thr Leu Leu Val Leu Ile Trp
35 40 45
Asn Asp His Ser Leu His Glu Pro Met Tyr Tyr Phe Leu Ala Met Leu
50 55 60
Ala Asp Thr Asp Leu Gly Met Thr Phe Thr Thr Met Pro Thr Val Leu
65 70 75 80
Gly Val Leu Leu Leu Asp Gln Arg Glu Ile Ala His Ala Ala Cys Phe
85 90 95
Thr Gln Ser Phe Ile His Ser Leu Ala Ile Val Glu Ser Gly Ile Leu
100 105 110
Leu Val Leu Ala Tyr Asp Cys Phe Ile Ala Ile Arg Thr Pro Leu Arg
115 120 125
Tyr Asn Cys Ile Leu Thr Asn Ser Arg Val Met Asn Ile Gly Leu Gly
130 135 140
Val Leu Met Arg Gly Phe Met Ser Ile Leu Pro Ile Ile Leu Ser Leu
145 150 155 160
Tyr Cys Tyr Pro Tyr Cys Gly Ser Arg Ala Leu Leu His Thr Phe Cys
165 170 175
Leu His Gln Asp Val Ile Lys Leu Ala Cys Ala Asp Ile Thr Phe Asn
180 185 190
His Ile Tyr Pro Ile Ile Gln Thr Ser Leu Thr Val Phe Leu Asp Ala
195 200 205
Leu Ile Ile Ile Phe Ser Tyr Ile Leu Ile Leu Lys Thr Val Met Gly
210 215 220
Ile Ala Ser Gly Gln Glu Glu Ala Lys Ser Leu Asn Thr Cys Val Ser
225 230 235 240
His Ile Ser Cys Val Leu Val Phe His Ile Thr Val Met Gly Leu Ser
245 250 255
Phe Ile His Arg Phe Gly Lys His Ala Pro His Val Val Pro Ile Thr
260 265 270
Met Ser Tyr Val His Phe Leu Phe Pro Pro Phe Val Asn Pro Ile Ile
275 280 285
Tyr Ser Ile Lys Thr Lys Gln Ile Gln Arg Ser Ile Ile Arg Leu Phe
290 295 300
Ser Gly Gln Ser Arg Ala
305 310
<210> 18
<211> 966
<212> DNA
<213> Homo sapiens
<400> 18
atgtggtcca attacagtgc tgctcccttc ttactgactg gcttcctggg ctcagagaca 60
atacaccact ggatctctat tcccttcttt gttatctact tctccatcct ttttggaaat 120
ggtacacttc ttcttctcat ttggagtgac cacggtcttc atgagcccat gtactacttc 180
cttgccatgc tggcaggtac agacctggga atgacattaa caacaatgcc cacagtcctg 240
ggtgttctac tgctagacca cagggagatt gcccagagtg cctgtttcac tcagtcctac 300
ttcattcact cactggcttt tgtagaatca ggtgtcttac ttgccatggc ctatgaccgt 360
tttattgcca tccgtgcccc tcttaggtac aactcaattc ttaccaactc tcgagtgatg 420
aagataggac ttggagtact gatgaggggc tttgtgtccc ttgtgccccc aattctacct 480
ctctattggt tcccatattg tcgttctcat gttctttccc atgctttttg cctccaccag 540
gatgtcatga aactcgcctg tgctgatata acatttaacc gcatataccc agtggttctg 600
gttgctttga ctttcttcct agatgttctg atcatcatct tttcttatat cttgatcctt 660
aagacagtta tgggcattgc ctctggagag gagcgagcaa aggctctcaa cacatgtgtc 720
tcccatatta gctctgtcct ggtattttat atcactgtga ttggcctgag cttcatccat 780
aggtttggga aacatgtgcc acatgtggtc cacattacca tgagctatgt ctactttctc 840
tttcctccat tcatgaaccc cattatatac agcatcaaga cgaaacagat tcagaaaagc 900
atcattcgtt tattttctgg gcaaaataga actttcaaca ttattataaa aattaaattt 960
tcatga 966
<210> 19
<211> 312
<212> PRT
<213> Homo sapiens
<400> 19
Met Ser Ser Ser Gly Ser Ser His Pro Phe Leu Leu Thr Gly Phe Pro
1 5 10 15
Gly Leu Glu Glu Ala His His Trp Ile Ser Val Phe Phe Leu Phe Met
20 25 30
Tyr Ile Ser Ile Leu Phe Gly Asn Gly Thr Leu Leu Leu Leu Ile Lys
35 40 45
Glu Asp His Asn Leu His Glu Pro Met Tyr Phe Phe Leu Ala Met Leu
50 55 60
Ala Ala Thr Asp Leu Gly Leu Ala Leu Thr Thr Met Pro Thr Val Leu
65 70 75 80
Gly Val Leu Trp Leu Asp His Arg Glu Ile Gly Ser Ala Ala Cys Phe
85 90 95
Ser Gln Ala Tyr Phe Ile His Ser Leu Ser Phe Leu Glu Ser Gly Ile
100 105 110
Leu Leu Ala Met Ala Tyr Asp Arg Phe Ile Ala Ile Cys Asn Pro Leu
115 120 125
Arg Tyr Thr Ser Val Leu Thr Asn Thr Arg Val Val Lys Ile Gly Leu
130 135 140
Gly Val Leu Met Arg Gly Phe Val Ser Val Val Pro Pro Ile Arg Pro
145 150 155 160
Leu Tyr Phe Phe Leu Tyr Cys His Ser His Val Leu Ser His Ala Phe
165 170 175
Cys Leu His Gln Asp Val Ile Lys Leu Ala Cys Ala Asp Thr Thr Phe
180 185 190
Asn Arg Leu Tyr Pro Ala Val Leu Val Val Phe Ile Phe Val Leu Asp
195 200 205
Tyr Leu Ile Ile Phe Ile Ser Tyr Val Leu Ile Leu Lys Thr Val Leu
210 215 220
Ser Ile Ala Ser Arg Glu Glu Arg Ala Lys Ala Leu Ile Thr Cys Val
225 230 235 240
Ser His Ile Cys Cys Val Leu Val Phe Tyr Val Thr Val Ile Gly Leu
245 250 255
Ser Leu Ile His Arg Phe Gly Lys Gln Val Pro His Ile Val His Leu
260 265 270
Ile Met Ser Tyr Ala Tyr Phe Leu Phe Pro Pro Leu Met Asn Pro Ile
275 280 285
Thr Tyr Ser Val Lys Thr Lys Gln Ile Gln Asn Ala Ile Leu His Leu
290 295 300
Phe Thr Thr His Arg Ile Gly Thr
305 310
<210> 20
<211> 1454
<212> DNA
<213> Homo sapiens
<400> 20
ctctttttgc ctgctgccat tccagtagga tgtgacttgc ttctcctggc catggttgtg 60
aggcctcccc agctatgtgg aactatgttg agaaggaaca acatgatcta agtctttaga 120
aaccaagagc cacgactagc tgaggacaca agaattttca caaaacaaag tagagtgatt 180
ccatagatca ctattatagc tgcattacca gtcctaggac agaatatgga tgctgaaaaa 240
tgattctttc aaagtgctga gttgttgcct cacgcccagt tgtctggcat ctcttcactg 300
aaactattac agttagatct catctctatg ggtggaagga aaaaattctg agaatgatat 360
agtcgaagtg ataccttgaa ggtagaactg tgcactgtgg taataacaga aacagaaaga 420
catttcttct atataaaccc aggatgtcgt ccagcggcag ctcccatccc ttcctattga 480
ctggttttcc aggcttggag gaagctcatc actggatttc cgtatttttc ttgttcatgt 540
atatatccat cctttttggc aatggcaccc tccttcttct cattaaggaa gatcacaatc 600
ttcatgagcc catgtacttc tttctggcca tgctggctgc cacagacctg gggctggccc 660
tgaccacaat gcccacggtg ctgggagtcc tctggctgga tcacagggag attggaagtg 720
cggcctgctt ttcccaggcc tactttatac actcactttc ctttctcgag tctggcattc 780
tgcttgccat ggcctatgac cgttttattg ccatctgcaa ccctcttaga tatacctctg 840
tacttactaa tactcgagta gtgaagattg ggctgggagt tctgatgagg ggatttgtat 900
ccgttgttcc cccaatcagg cccctctatt tttttctgta ttgtcactcc catgttcttt 960
cacatgcatt ctgccttcac caggatgtca ttaaactcgc ctgtgctgat accaccttca 1020
accgactgta cccagctgtg cttgtagtct ttatatttgt gctggattat ctgattatct 1080
tcatctccta tgtgttgata ctcaagactg tcctgagcat tgcctccaga gaggagaggg 1140
ccaaggctct cattacctgt gtctcccata tctgctgtgt cctggttttt tatgtcacag 1200
tgattggatt gtctctgatt catcgttttg gaaagcaggt tccacatatt gttcacctca 1260
ttatgagcta tgcctatttt ctgttccctc cactaatgaa tcctataaca tatagtgtca 1320
agaccaagca gattcagaat gccattcttc acctttttac tacccataga attggaacct 1380
gatctccaat catcacagtc actctcacag aatgtaaaaa ctctaatatt tagcaggaga 1440
gcaaagaagt ctca 1454
<210> 21
<211> 324
<212> PRT
<213> Homo sapiens
<400> 21
Met Leu Gly Pro Ala Tyr Asn His Thr Met Glu Thr Pro Ala Ser Phe
1 5 10 15
Leu Leu Val Gly Ile Pro Gly Leu Gln Ser Ser His Leu Trp Leu Ala
20 25 30
Ile Ser Leu Ser Ala Met Tyr Ile Thr Ala Leu Leu Gly Asn Thr Leu
35 40 45
Ile Val Thr Ala Ile Trp Met Asp Ser Thr Arg His Glu Pro Met Tyr
50 55 60
Cys Phe Leu Cys Val Leu Ala Ala Val Asp Ile Val Met Ala Ser Ser
65 70 75 80
Val Val Pro Lys Met Val Ser Ile Phe Cys Ser Gly Asp Ser Ser Ile
85 90 95
Ser Phe Ser Ala Cys Phe Thr Gln Met Phe Phe Val His Leu Ala Thr
100 105 110
Ala Val Glu Thr Gly Leu Leu Leu Thr Met Ala Phe Asp Arg Tyr Val
115 120 125
Ala Ile Cys Lys Pro Leu His Tyr Lys Arg Ile Leu Thr Pro Gln Val
130 135 140
Met Leu Gly Met Ser Met Ala Val Thr Ile Arg Ala Val Thr Phe Met
145 150 155 160
Thr Pro Leu Ser Trp Met Met Asn His Leu Pro Phe Cys Gly Ser Asn
165 170 175
Val Val Val His Ser Tyr Cys Lys His Ile Ala Leu Ala Arg Leu Ala
180 185 190
Cys Ala Asp Pro Val Pro Ser Ser Leu Tyr Ser Leu Ile Gly Ser Ser
195 200 205
Leu Met Val Gly Ser Asp Val Ala Phe Ile Ala Ala Ser Tyr Ile Leu
210 215 220
Ile Leu Arg Ala Val Phe Asp Leu Ser Ser Lys Thr Ala Gln Leu Lys
225 230 235 240
Ala Leu Ser Thr Cys Gly Ser His Val Gly Val Met Ala Leu Tyr Tyr
245 250 255
Leu Pro Gly Met Ala Ser Ile Tyr Ala Ala Trp Leu Gly Gln Asp Ile
260 265 270
Val Pro Leu His Thr Gln Val Leu Leu Ala Asp Leu Tyr Val Ile Ile
275 280 285
Pro Ala Thr Leu Asn Pro Ile Ile Tyr Gly Met Arg Thr Lys Gln Leu
290 295 300
Leu Glu Gly Ile Trp Ser Tyr Leu Met His Phe Leu Phe Asp His Ser
305 310 315 320
Asn Leu Gly Ser
<210> 22
<211> 975
<212> DNA
<213> Homo sapiens
<400> 22
atgctgggtc cagcttacaa ccacacaatg gaaacccctg cctccttcct ccttgtgggt 60
atcccaggac tgcaatcttc acatctttgg ctggctatct cactgagtgc catgtacatc 120
acagccctgt taggaaacac cctcatcgtg actgcaatct ggatggattc cactcggcat 180
gagcccatgt attgctttct gtgtgttctg gctgctgtgg acattgttat ggcctcctcc 240
gtggtaccca agatggtgag catcttctgc tcgggagaca gctccatcag ctttagtgct 300
tgtttcactc agatgttttt tgtccactta gccacagctg tggagacggg gctgctgctg 360
accatggctt ttgaccgcta tgtagccatc tgcaagcctc tacactacaa gagaattctc 420
acgcctcaag tgatgctggg aatgagtatg gccgtcacca tcagagctgt cacattcatg 480
actccactga gttggatgat gaatcatcta cctttctgtg gctccaatgt ggttgtccac 540
tcctactgta agcacatagc tttggccagg ttagcatgtg ctgaccccgt gcccagcagt 600
ctctacagtc tgattggttc ctctcttatg gtgggctctg atgtggcctt cattgctgcc 660
tcctatatct taattctcag ggcagtattt gatctctcct caaagactgc tcagttgaaa 720
gcattaagca catgtggctc ccatgtgggg gttatggctt tgtactatct acctgggatg 780
gcatccatct atgcggcctg gttggggcag gatatagtgc ccttgcacac ccaagtgctg 840
ctagctgacc tgtacgtgat catcccagcc actttaaatc ccatcatcta tggcatgagg 900
accaaacaat tgctggaggg aatatggagt tatctgatgc acttcctctt tgaccactcc 960
aacctgggtt catga 975
<210> 23
<211> 318
<212> PRT
<213> Homo sapiens
<400> 23
Met Ser Asp Ser Asn Leu Ser Asp Asn His Leu Pro Asp Thr Phe Phe
1 5 10 15
Leu Thr Gly Ile Pro Gly Leu Glu Ala Ala His Phe Trp Ile Ala Ile
20 25 30
Pro Phe Cys Ala Met Tyr Leu Val Ala Leu Val Gly Asn Ala Ala Leu
35 40 45
Ile Leu Val Ile Ala Met Asp Asn Ala Leu His Ala Pro Met Tyr Leu
50 55 60
Phe Leu Cys Leu Leu Ser Leu Thr Asp Leu Ala Leu Ser Ser Thr Thr
65 70 75 80
Val Pro Lys Met Leu Ala Ile Leu Trp Leu His Ala Gly Glu Ile Ser
85 90 95
Phe Gly Gly Cys Leu Ala Gln Met Phe Cys Val His Ser Ile Tyr Ala
100 105 110
Leu Glu Ser Ser Ile Leu Leu Ala Met Ala Phe Asp Arg Tyr Val Ala
115 120 125
Ile Cys Asn Pro Leu Arg Tyr Thr Thr Ile Leu Asn His Ala Val Ile
130 135 140
Gly Arg Ile Gly Phe Val Gly Leu Phe Arg Ser Val Ala Ile Val Ser
145 150 155 160
Pro Phe Ile Phe Leu Leu Arg Arg Leu Pro Tyr Cys Gly His Arg Val
165 170 175
Met Thr His Thr Tyr Cys Glu His Met Gly Ile Ala Arg Leu Ala Cys
180 185 190
Ala Asn Ile Thr Val Asn Ile Val Tyr Gly Leu Thr Val Ala Leu Leu
195 200 205
Ala Met Gly Leu Asp Ser Ile Leu Ile Ala Ile Ser Tyr Gly Phe Ile
210 215 220
Leu His Ala Val Phe His Leu Pro Ser His Asp Ala Gln His Lys Ala
225 230 235 240
Leu Ser Thr Cys Gly Ser His Ile Gly Ile Ile Leu Val Phe Tyr Ile
245 250 255
Pro Ala Phe Phe Ser Phe Leu Thr His Arg Phe Gly His His Glu Val
260 265 270
Pro Lys His Val His Ile Phe Leu Ala Asn Leu Tyr Val Leu Val Pro
275 280 285
Pro Val Leu Asn Pro Ile Leu Tyr Gly Ala Arg Thr Lys Glu Ile Arg
290 295 300
Ser Arg Leu Leu Lys Leu Leu His Leu Gly Lys Thr Ser Ile
305 310 315
<210> 24
<211> 1065
<212> DNA
<213> Homo sapiens
<400> 24
agcccttcat ttgccaggaa gaatgtcaga ttccaacctc agtgataacc atcttccaga 60
caccttcttc ttaacaggga tcccagggct ggaggctgcc cacttctgga ttgccatccc 120
tttctgtgcc atgtatcttg tagcactggt tggaaatgct gccctcatcc tggtcattgc 180
catggacaat gctcttcatg cacctatgta cctcttcctc tgccttctct cactcacaga 240
cctggctctc agttctacca ctgtgcccaa gatgctggcc attttgtggc tccatgctgg 300
tgagatttcc tttggtggat gcctggccca gatgttttgt gtccattcta tctatgctct 360
ggagtcctca attctacttg ccatggcctt tgataggtat gtggctatct gtaacccatt 420
aaggtacaca accattctca accatgctgt cataggcaga attggctttg ttgggctatt 480
ccgtagtgtg gctattgtct cccccttcat cttcttgctg aggcgactcc cctactgtgg 540
tcaccgtgtc atgacacaca catactgtga gcatatgggc attgcccgac tggcctgtgc 600
caacatcact gtcaatattg tctatgggct aactgtggct ctgctggcca tgggactgga 660
ttccattctc attgccattt cctatggctt tatcctccat gcagtctttc atcttccatc 720
tcatgatgcc cagcacaaag ctctgagtac ctgtggctcc cacattggca tcatcctggt 780
tttctacatc cctgccttct tctccttcct cacccaccgc tttggtcacc acgaagtccc 840
caagcatgtg cacatctttc tggctaatct ctatgtgctg gtgcctcctg tactcaatcc 900
tattctctat ggagctagaa ccaaggagat tcggagtcga cttctaaaac tgcttcacct 960
ggggaagact tcaatatgaa tgctgagcag aagttggaga tttaaaaaaa agtgtaggat 1020
ggctgtctag atacatttac atggacacac agtgatgatg tgaaa 1065
<210> 25
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> COL1A1 F primer
<400> 25
acatgttcag ctttgtggac c 21
<210> 26
<211> 20
<212> DNA
<213> Artificial Sequence
<220>
<223> COL1A1 R primer
<400> 26
tgtacgcagg tgattggtgg 20
<210> 27
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> OR1F2P F primer
<400> 27
ctgagggtcc catccacaaa g 21
<210> 28
<211> 25
<212> DNA
<213> Artificial Sequence
<220>
<223> OR1F2P R primer
<400> 28
gggaggatga agttctgaaa tatgg 25
<210> 29
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2A9P F primer
<400> 29
catcctgagg agcagcagaa g 21
<210> 30
<211> 23
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2A9P R primer
<400> 30
aggcaataat tgggcatcaa gag 23
<210> 31
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2A20P F primer
<400> 31
tccctgttct atgtcttcac cc 22
<210> 32
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2A20P R primer
<400> 32
caggaggcat tcagtatgtg c 21
<210> 33
<211> 23
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2AK2 F primer
<400> 33
tctatgcaac cactctcttt acc 23
<210> 34
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2AK2 R primer
<400> 34
aaatactgcc accgccttat cc 22
<210> 35
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> OR10A4 F primer
<400> 35
cccttgcact acccagtcat c 21
<210> 36
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> OR10A4 R primer
<400> 36
tcagagcctc cagttcaaac ac 22
<210> 37
<211> 19
<212> DNA
<213> Artificial Sequence
<220>
<223> OR10A5 F primer
<400> 37
caaaggacac gggccaaac 19
<210> 38
<211> 23
<212> DNA
<213> Artificial Sequence
<220>
<223> OR10A5 R primer
<400> 38
cagcaatgcg agtataggaa cac 23
<210> 39
<211> 19
<212> DNA
<213> Artificial Sequence
<220>
<223> GAPDH F primer
<400> 39
aaatcaagtg gggcgatgc 19
<210> 40
<211> 20
<212> DNA
<213> Artificial Sequence
<220>
<223> GAPDH R primer
<400> 40
agggggcaga gatgatgacc 20
<210> 41
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> OR7E12P F primer
<400> 41
ttgctgagtg tggattgctc t 21
<210> 42
<211> 20
<212> DNA
<213> Artificial Sequence
<220>
<223> OR7E12P R primer
<400> 42
tgatgagtag gttccccagc 20
<210> 43
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2A1/2A42 F primer
<400> 43
gcatgacgca gacctttctc t 21
<210> 44
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2A1/2A42 R primer
<400> 44
gcagtcttag gatgagaacc a 21
<210> 45
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2A7 F primer
<400> 45
gacctcattc aacctcacaa tc 22
<210> 46
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2A7 R primer
<400> 46
atttcattca gcggcaaaca c 21
<210> 47
<211> 20
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2AE1 F primer
<400> 47
tgtggcagaa gaatcagacc 20
<210> 48
<211> 20
<212> DNA
<213> Artificial Sequence
<220>
<223> OR2AE1 R primer
<400> 48
gagaatggtg agggtgttgc 20
<210> 49
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> OR51B4 F primer
<400> 49
ctgctagacc agagggagat tg 22
<210> 50
<211> 25
<212> DNA
<213> Artificial Sequence
<220>
<223> OR51B4 R primer
<400> 50
tgttcatcac tcgggaattg gtaag 25
<210> 51
<211> 26
<212> DNA
<213> Artificial Sequence
<220>
<223> OR51B5 F primer
<400> 51
actgtggtaa taacagaaac agaaag 26
<210> 52
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> OR51B5 R primer
<400> 52
accagtcaat aggaagggat gg 22
<210> 53
<211> 23
<212> DNA
<213> Artificial Sequence
<220>
<223> OR51I1 F primer
<400> 53
ccatgctcaa ccctatcatc tac 23
<210> 54
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> OR51I1 R primer
<400> 54
ggcctgggat ttatggaaga ac 22
<210> 55
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> OR52D1 F primer
<400> 55
tctccccctt catcttcttg ct 22
<210> 56
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> OR52D1 R primer
<400> 56
gagccacagt tagcccatag a 21
Claims (17)
- OR1F2P, OR2A9P, OR2A20P, OR2AK2, OR10A4, OR10A5, OR7E12P, OR2A1/2A42, OR2A7, OR2AE1, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량을 측정하는 제제를 유효성분으로 포함하는 피부 노화의 진단용 조성물.
- 제 1 항에 있어서, 상기 피부 노화는 스트레스로 인한 피부 노화인 것을 특징으로 하는 조성물.
- 제 1 항에 있어서, 상기 조성물은 OR1F2P, OR2A9P, OR2A20P, OR2AK2, OR10A4 및 OR10A5로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량을 측정하는 제제를 포함하는 것을 특징으로 하는 조성물.
- 제 3 항에 있어서, 상기 조성물은 자외선 노출에 의한 피부의 노화 정도를 예측하는 것을 특징으로 하는 조성물.
- 제 4 항에 있어서, 상기 자외선 노출에 의한 피부의 노화는 미세주름, 과색소침착, 피부 탄력의 소실 및 수분 결핍으로 구성된 군으로부터 선택되는 하나 이상의 광노화(Photoaging) 현상인 것을 특징으로 하는 조성물.
- 제 1 항에 있어서, 상기 조성물은 OR7E12P 유전자 또는 이의 인코딩 단백질의 발현량을 측정하는 제제를 포함하는 것을 특징으로 하는 조성물.
- 제 6 항에 있어서, 상기 조성물은 염증에 의한 피부의 노화 정도를 예측하는 것을 특징으로 하는 조성물.
- 제 7 항에 있어서, 상기 염증에 의한 피부의 노화는 아토피 피부염, 피부근염, 피부경화증, 천포창, 환상홍반 및 홍반루푸스로 구성된 군으로부터 선택되는 염증성 또는 자가면역성 피부질환으로 인한 피부 노화인 것을 특징으로 하는 조성물.
- 제 1 항에 있어서, 상기 조성물은 OR2A1/2A42, OR2A7, OR2AE1, OR2AK2, OR10A4, OR10A5, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량을 측정하는 제제를 포함하는 것을 특징으로 하는 조성물.
- 제 9 항에 있어서, 상기 조성물은 피부의 장벽 기능 손상에 의한 피부의 노화 정도를 예측하는 것을 특징으로 하는 조성물.
- 제 1 항에 있어서, 상기 유전자의 발현량을 측정하는 제제는 상기 유전자의 핵산 분자에 특이적으로 결합하는 프라이머 또는 프로브인 것을 특징으로 하는 조성물.
- 제 1 항에 있어서, 상기 단백질의 발현량을 측정하는 제제는 상기 단백질에 특이적으로 결합하는 항체 또는 이의 항원 결합 단편; 또는 상기 단백질에 특이적으로 결합하는 앱타머인 것을 특징으로 하는 조성물.
- 개체로부터 분리된 생물학적 시료 내에서 OR1F2P, OR2A9P, OR2A20P, OR2AK2, OR10A4, OR10A5, OR7E12P, OR2A1/2A42, OR2A7, OR2AE1, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량을 측정하는 단계를 포함하는 피부 노화의 진단에 필요한 정보를 제공하는 방법.
- 제 13 항에 있어서, 상기 생물학적 시료는 피부조직 또는 피부조직 유래 세포를 포함하는 것을 특징으로 하는 방법.
- 제 14 항에 있어서, 상기 피부조직 유래 세포는 피부 섬유아세포(dermal fibroblast)인 것을 특징으로 하는 방법.
- 다음의 단계를 포함하는 피부 노화 개선용 조성물의 스크리닝 방법:
(a) OR1F2P, OR2A9P, OR2A20P, OR2AK2, OR10A4, OR10A5, OR7E12P, OR2A1/2A42, OR2A7, OR2AE1, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자를 발현하는 세포를 포함하는 생물학적 시료에 후보물질을 접촉시키는 단계;
(b) 상기 시료 내 상기 유전자 또는 상기 유전자가 인코딩하는 단백질의 발현량 또는 활성을 측정하는 단계,
상기 OR1F2P 유전자 또는 이의 인코딩하는 단백질의 발현량 또는 활성이 감소하거나, 상기 OR2A9P, OR2A20P, OR2AK2, OR10A4, OR10A5, OR7E12P, OR2A1/2A42, OR2A7, OR2AE1, OR51B4, OR51B5, OR51I1 및 OR52D1로 구성된 군으로부터 선택되는 하나 이상의 유전자 또는 이들이 인코딩하는 단백질의 발현량 또는 활성이 증가한 경우, 상기 후보물질은 피부 노화 개선용 조성물로 판정한다.
- 제 16 항에 있어서, 상기 세포는 피부 섬유아세포(dermal fibroblast)인 것을 특징으로 하는 방법.
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PCT/KR2022/000538 WO2022154462A1 (ko) | 2021-01-12 | 2022-01-12 | 피부 섬유아세포에서 발현하는 노화 관련 후각 수용체 유전자 및 이들의 용도 |
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US10209239B1 (en) * | 2017-08-16 | 2019-02-19 | Aromyx Corporation | Method of making an aromagraph comprising ectopic olfactory receptors |
KR102032540B1 (ko) * | 2018-01-03 | 2019-10-15 | 의료법인 성광의료재단 | 피부노화 특이적 후성유전자 마커 및 이를 이용한 피부노화의 탐지방법 |
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Title |
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비특허문헌 1. Romain Duroux et al., A Rose Extract Protects the Skin against Stress Mediators: A Potential Role of Olfactory Receptors. Molecules 25:4743(2020) |
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