Molecular Genetics of the ET-1/AngII Receptor

描述(申请人提供)：内皮素-1(ET-1)和血管紧张素II 血管紧张素转换酶(AngII)是心脏正常运转所必需的关键荷尔蒙 心血管系统，既有中枢作用，也有外围作用。两种荷尔蒙 系统与心血管疾病--高血压， 内皮功能障碍、心肌肥大和衰竭。几种同工受体 最近对ET-1和AngII进行了鉴定。在这些人中，双重的 ET-1/AngII受体，我们通过一种基于和支持ET-1/AngII受体的方法分离出来 分子识别理论阐明了新的信息，为 寻找新的假设和问题。这项提议的长期目标是 剖析ET-L/AngII受体基因对血液的特异性贡献 正常和病理生理状态下的压力调节，如高血压。 在本研究方案中，我们将检验以下假设：1)基于 我们的初步数据显示：1)ET-II的性别偏见 /AngII受体基因座与高血压相关的F_2(S×R)杂交组合， 和b)Dahl盐敏感和Dahl之间的结构差异 耐盐大鼠ET-1/AngII受体，我们假设ET-1/AngII受体 受体以性别特有的方式影响血压。 Dahl大鼠模型。2)根据检测到的ET-L/AngII受体的丰度 脑和心血管组织中的mRNA，我们假设该受体是 ET-1和AngII在这些器官中的一个突出的生理靶点，以及 因此在血压调节中起着重要的作用。调查 以下具体目标将检验这些假设：1.性别特定的角色 ET-1/AngII受体与盐敏感型高血压易感性的关系 1a)Dahl-S和Dahl R-1/AngII受体的功能特性 它们有两种不同的氨基酸。1b)三个自交系的建立 携带外源性Dahl R ET-1/AngII受体的Dahl S大鼠系 TG[RET-1/AngIIR]和三个对照Dahl S大鼠系(外源Dahl S) ET-1/Angil受体，Tg[Set-1/AnglIr]，由同源Dahl R 5‘驱动 侧翼监管区域。1c)确定转基因分子表型， 即外源(转基因)Dahl R和Dahl S等的相对量。 血管紧张素-1受体和内源性血管紧张素-1受体的表达 甘油三酯[RET-1/AngIIr]和甘油三酯[SET-1/AngIIr]大鼠的水平。1D) 测定Dahl R和Dahl S ET-1/AngII的生理效应 年龄匹配的非转基因受体转基因差异分析 近交系S大鼠(雌雄)及转基因近交系TG[Ret-1/AnglIr] 和甘油三酯-S大鼠系：1)发病、病程和 高血压程度；2)寿命；3)高血压肾病严重程度。 2.对ET-1/AngII受体生理作用的剖析(S)。2a)组织 特定的、空间和时间的表达模式在儿童发育中的作用 ET-1/AngII受体基因。2b)靶向干扰ET-1/Angil受体 “基因敲除”小鼠模型的研究进展 生理学在综合生物实验系统中的作用。我们的建议 研究将阐明ET-1/AngII受体的生理作用 在整个发展过程中。此外，它还将展示特定性别的 ET-1/AngII受体在盐敏感性高血压中的作用 Dahl大鼠模型，从而提供了一种有效的优先方案来调查 该受体在人类原发性高血压中的潜在作用。

DESCRIPTION (provided by the applicant): Endothelin-1 (ET-1) and angiotensin II (AngII) are critical hormones necessary for the appropriate function of the cardiovascular system, acting centrally as well as peripherally. Both hormonal systems have been implicated in cardiovascular pathology -- hypertension, endothelial dysfunction, cardiac hypertrophy and failure. Several isoreceptors for both ET-1 and AngII have been recently characterized. Of these, the dual ET-1/AngII receptor, isolated by us via an approach based on and supporting the molecular recognition theory elucidates novel information providing the bases for new hypotheses and questions. The long-term objective of this proposal is to dissect the specific contribution of the ET-l/AngII receptor gene to blood pressure regulation in normal and pathophysiological states like hypertension. In this research proposal, we will test the following hypotheses: 1) Based on our preliminary data showing a) gender-bias o 1I the cosegregation of the ET-II /AngII receptor locus with hypertension in an F2 (Dahl S x Dahl R) intercross, and b) structural differences between the Dahl salt-sensitive and the Dahl salt-resistant rat ET-1/AngII receptors, we hypothesize that the ET-1/AngII receptor acts in a gender specific manner influencing blood pressure in the Dahl rat model. 2) Based on the detected abundance of the ET-l/AngII receptor mRNA in brain and cardiovascular tissues, we hypothesize that this receptor is a prominent physiologic target for ET- 1 and AngII in these organs, and therefore plays an important role in blood pressure regulation. Investigating the following specific aims will test these hypotheses: 1. Gender specific role of the ET-1/AngII receptor in susceptibility to salt-sensitive hypertension. 1A) Functional characterization of the Dahl S and Dahl R ET-1/AngII receptors that differ in two amino acids. 1B) Establishment of three inbred trans genie Dahl S rat lines bearing exogenous Dahl R ET-1/AngII receptor, Tg[RET-1/AngIIr], and three control Dahl S rat lines bearing exogenous Dahl S ET- 1/AngIl receptor, Tg[SET- 1/AnglIr], driven by the cognate Dahl R 5' flanking regulatory region. 1C) Determine the transgenic molecular phenotype, that is the relative amounts of exogenous (transgenic) Dahl R and Dahl S ET- 1/Angll receptor mRNA and endogenous Dahl S ET-1/AngIl receptor mRNA expression levels in Tg[RET-1/AngIIr] and Tg[SET-1/AngIIr} rats respectively. 1D) Determine the physiological effects of the Dahl R and Dahl S ET- 1/AngII receptor transgene by analyzing differences between age-matched non-transgenic inbred Dahl S rats (males and females) and transgenic inbred Tg[RET- 1/AnglIr] and Tg[SET1I AngIIr] Dahl S rat lines with respect to 1) the onset, course and degree of hypertension; 2) lifespan; 3) severity of hypertensive renal disease. 2. Dissection of physiologic role(s) of the ET-1/AngII receptor. 2A) Tissue specific, spatial and temporal expression patterns in development of the ET-1/AngII receptor gene. 2B) Targeted disruption of the ET- 1/AngIl receptor gene and development of "knockout" mouse model in order to define its physiologic role in an integrated biologic experimental system. Our proposed studies will elucidate the physiological role of the ET- 1 /AngII receptor throughout development. Furthermore, it will demonstrate the gender specific involvement of the ET- 1/AngII receptor in salt-sensitive hypertension in the Dahl rat model, thus providing a validated prioritization scheme to investigate the potential role of this receptor in human essential hypertension.