Gender-Dependent Regulation of Pitx2 in Atrial Fibrillation

项目总结 房颤是美国最常见的持续性心律失常，其终生风险为25%。 占所有心血管疾病的三分之一。与房颤相关的治疗和护理费用大致 仅在美国每年就有260亿美元，占所有医疗保险支出的10%。相关的令人衰弱的并发症 房颤包括心力衰竭和中风。有证据表明，房颤是大多数缺血性和隐源性中风的基础， 女性比男性有更高的风险。除了中风，房颤的几个危险因素、结果和治疗 展示对性别差异的机械理解是有限的。最常见的变种 非编码的4q25基因座与房颤的风险密切相关，这意味着成对的类基因的顺式调节元件 同源结构域转录因子，PITX2。为了了解房颤的分子驱动因素，有必要了解 了解PITX2的上游调控和下游行动。PITX2在左心房有表达 (LA)和肺静脉(PV)心肌。虽然研究表明PITX2在房颤中的作用 LA心肌，也被认为是起源于PV的房颤的基础，PV是最常见的触发因素 人类患者心律失常的部位。虽然Pitx2的动物模型对房颤易感，但直接的分子 由PITX2调控的机制仍未确定。利用Pitx2 CRISPR编辑的小鼠，Pitx2在LA中的作用 和PV心肌将被询问，同时解决潜在的性别差异的Pitx2调节。这个 核心假设是基因调控网络中的性别差异是差异的基础 房颤--男性和女性的易感性。为了解决这一假设，第一个目标是检查 Pitx2表达中性别偏见和房颤风险的转录基础。Motif分析确定了多个高度- 保守的激素受体结合位点，包括雄激素和雌激素受体，位于4q25相关 调节元件和Pitx2启动子。CRISPR介导的4q25相关调控基因的缺失 Element使男性特有的Pitx2减少，并增加了心脏起搏导致的房颤风险。因此，这个目标 会问性激素依赖的表达是否通过直接影响房颤负荷和Pitx2的表达 转录调控。在第二个目标中，将询问PITX2的下游目标以发现 不同性别的表观基因组差异是否影响LA和PV中的转录因子环境 心肌。尽管PITX2与房颤有很强的相关性，但PITX2在LA/PV心肌中的功能 人们对此知之甚少。此外，在房颤和其他疾病中，男性和女性之间存在显著差异 心血管疾病；然而，很少有心血管系统的表观遗传学或基因组研究 性别。这一目标旨在阐明PITX2在房颤中的作用，同时采取渐进的方法 在这项遗传和表观基因组研究中纳入和利用性别差异。总而言之， 拟议的项目旨在确定PITX2在房颤中的上游调控和下游作用 镜头固有的性别差异，目的是确定新的途径来治疗男性和女性的房颤。

PROJECT SUMMARY Atrial fibrillation (AF) is the most common sustained arrhythmia in the United States with a 25% lifetime risk, and accounts for one-third of all cardiovascular diseases. Treatment and care associated with AF costs roughly $26 billion/year in the US alone and accounts for 10% of all Medicare spending. Debilitating complications linked to AF include heart failure and stroke. Evidence suggests AF underlies most ischemic and cryptogenic strokes, with women at a higher risk than men. In addition to stroke, several risk factors, outcomes, and treatments of AF demonstrate gender differences for which there is limited mechanistic understanding. Common variation at the non-coding 4q25 locus is resoundingly linked to AF risk, implicating cis-regulatory elements of the paired-like homeodomain transcription factor, PITX2. To understand the molecular drivers of AF, it is imperative to understand both the upstream regulation and downstream actions of PITX2. PITX2 is expressed in the left atrial (LA) and pulmonary vein (PV) myocardium of humans and mice. While work has implicated PITX2 in AF in the LA myocardium, it has also been suggested to underlie AF originating from the PV, the most common trigger site for arrhythmia in human patients. Although animal models of Pitx2 are susceptible to AF, the direct molecular mechanisms regulated by Pitx2 remain undetermined. Using Pitx2 CRISPR-edited mice, the role of Pitx2 in LA and PV myocardium will be interrogated while addressing underlying gender differences in Pitx2 regulation. The central hypothesis is that gender-specific differences in gene regulatory networks underlie differences in AF-susceptibility in men and women. To address this hypothesis, the first aim proposes to examine the transcriptional basis for gender bias in Pitx2 expression and AF-risk. Motif analysis identifies multiple, highly- conserved hormone-receptor binding sites, including androgen- and estrogen-receptor, at a 4q25-associated regulatory element and the Pitx2 promoter. CRISPR-mediated deletion of the 4q25-associated regulatory element confers male-specific reduction in Pitx2 and increased risk of AF by cardiac pacing. Therefore, this aim will ask whether sex-hormone dependent expression influence AF burden and Pitx2 expression through direct transcriptional regulation. In the second aim, the downstream targets of PITX2 will be interrogated to discover whether epigenomic differences between the genders influence the transcription factor milieu in the LA and PV myocardium. Despite the strong association between PITX2 and AF, the function of PITX2 in LA/PV myocardium is poorly understood. Furthermore, significant differences between men and women are present in AF and other cardiovascular diseases; however, few epigenetic or genomic studies of the cardiovascular system examine gender. This aim is designed to illuminate the role of PITX2 in AF while taking a progressive approach to incorporating and leveraging gender differences in this genetic and epigenomic investigation. Altogether, the proposed project aims to identify the upstream regulation and downstream actions of PITX2 in AF through the lens of inherent gender differences with the goal of identifying novel avenues to treat AF in men and women.