Functional characterization of ATP1A1 and DEspR variants associated with essentia

描述（由申请人提供）：项目总结原发性高血压是一个主要的公共卫生问题，因为它的患病率高，并且是发达国家主要死亡和发病原因的主要风险因素-冠状动脉疾病、中风、慢性肾脏疾病和外周血管疾病。阐明潜在的遗传机制是至关重要的，但仍然难以捉摸，由于其多基因的性质和环境因素带来的复杂性。我们报道了来自撒丁岛北方的病例对照高血压队列中ATP 1A 1（α 1 Na，K-ATP酶，P<0.000005）和DEspR（内皮素1/血管生长因子信号肽双重受体，P<0.03）与高血压/正常血压的关系。我们检测到这两个基因座与高血压/正常血压的男性比女性更强的关联，指出5 '调控区作为其关联的潜在变异体。后续研究发现，ATP 1A 1和DEspR启动子区域内的4 T缺失/插入多态性（4 Tdelins）和C/T（rs6535847）多态性分别与男性人群中高血压易感性降低相关。C/T（rs6535847）多态性修饰了DEspR启动子区的一个CATAAAA序列，产生了一个新的TATAAAA- box，提示可能对DEspR转录有影响。这些结果构成了我们目前应用的基础，我们计划研究ATP 1A 1 4 T ins和DEspR rs6535847 T等位基因作为潜在的功能变体，解释这两种保护性等位基因在男性撒丁岛人群中赋予的高血压易感性降低的风险。因此，下列具体目标被列为优先事项：我们将检测携带4 T ins（p4 Tins）或4 T del（p4 Tdel）等位基因的ATP 1A 1启动子区和携带rs6535847 C使用SEAP（人胎盘碱性磷酸酶的分泌形式）作为报告分子监测组织培养细胞中的rs6535847 T等位基因（pCATAAAA）或rs6535847 T等位基因（pTATAAAA）的活性。目标2：我们将通过无线电遥测测量年轻（4月龄）和老龄（16月龄）DEspR、ATP 1A 1和野生型小鼠的血压，研究DEspR和ATP 1A 1单倍不足对血压的影响。这将在生物学背景下评估不同DEspR和ATP 1A 1表达水平对血压的推定影响。实现提出的具体目标将阐明DEspR和ATP 1A 1功能多态性，赋予性别特异性保护原发性高血压的遗传隔离人口从北方撒丁岛。此外，这些结果可以为普通人群中高血压遗传易感性的研究方向提供关键的见解。公共卫生相关性：项目叙述高血压是心脏病、中风和肾衰竭的主要危险因素。尽管对高血压及其靶器官相关并发症的遗传机制的研究越来越多，但高血压的遗传基础仍有待充分阐明。我们的研究将有助于建立两个遗传因素与发展的高血压在北方撒丁岛人口。这一信息将提供一个框架，进一步调查各自的作用，在一般人群中的高血压，并有助于改善干预和预防策略，原发性高血压及其靶器官并发症。

DESCRIPTION (provided by applicant): Project Summary Essential hypertension is a major public health concern due to its high prevalence and its role as a leading risk factor for leading causes of death and morbidity in the developed world - coronary artery disease, stroke, chronic renal disease and peripheral vascular disease. Elucidation of underlying genetic mechanism is critical but remains elusive due to its polygenic nature and added complexity brought on by environmental factors. We reported the association of ATP1A1 (alpha 1 Na,K-ATPase, P<0.000005) and DEspR (dual endothelin-1/vascular growth factor signal peptide receptor, P<0.03) with hypertension/normotension in a case-control hypertension cohort from northern Sardinia. We detected stronger association of both loci with hypertension/normotension in males than in females pointing to the 5'-regulatory region as harboring putative variants underlying their associations. Follow up studies identified a 4T deletion/insertion polymorphism (4Tdelins) and a C/T (rs6535847) polymorphism within the ATP1A1 and DEspR promoter regions respectively associated with decreased susceptibility to hypertension in the male population. The C/T (rs6535847) polymorphism modifies a CATAAAA sequence present in DEspR promoter region to generate a new TATAAAA- box, suggesting a putative effect on DEspR transcription. These results form the basis for our current application in which we plan to investigate the ATP1A1 4T ins and DEspR rs6535847 T alleles as potential functional variants accounting for the decreased risk of hypertension susceptibility conferred by these two protective alleles in the male Sardinian population. Thus, the following specific aims are prioritized: AIM 1: We will test the transcriptional activity of ATP1A1 promoter region carrying either the 4T ins (p4Tins) or the 4T del (p4Tdel) alleles and DEspR promoter region harboring either the rs6535847 C (pCATAAAA) or the rs6535847 T alleles (pTATAAAA) in tissue culture cells using SEAP (a secreted form of human placental alkaline phosphatase) as a reporter molecule to monitor activity. AIM 2: We will investigate the effect of DEspR and ATP1A1 haploinsufficiency on blood pressure by measuring blood pressure by radiotelemetry in young (4 months of age) and aging (16 months of age) DEspR, ATP1A1 and wild-type mice. This will assess in a biological context the putative effects of differential DEspR and ATP1A1 expression levels on blood pressure. Accomplishing the proposed specific aims will elucidate DEspR and ATP1A1 functional polymorphisms that confer sex- specific protection to essential hypertension in a genetically isolated population from northern Sardinia. Moreover, these results could shed key insight into setting the direction for the investigation of genetic susceptibility to hypertension in the general population. PUBLIC HEALTH RELEVANCE: Project Narrative Hypertension is a leading risk factor for heart disease, stroke and renal failure. Despite increasing efforts to decipher the genetic mechanisms of hypertension and its target organ associated complications, the genetic underpinnings of hypertension remain to be fully elucidated. Our research will help to establish two genetic factors associated with development of hypertension in a northern Sardinian population. This information will provide a framework to investigate further their respective roles in hypertension in the general population and help to improve intervention and prevention strategies for essential hypertension and its target organ complications.