Investigation of alternative splicing in response to low and disturbed flow

 描述（由申请人提供）：项目摘要 我的长期目标是在一个顶级学术机构领导一个研究小组，追求我的兴趣：血管发育和疾病的遗传调控，重点是内皮和血液动力学激活。我对血流对血管系统的影响非常着迷，在过去的10年里，我一直专注于内皮功能的转录调控。我相信体内遗传学和成像将是理解内皮细胞、血流和募集的血细胞之间复杂相互作用的关键。反过来，这对于理解血管发育和疾病（如动脉瘤和动脉粥样硬化）至关重要，这些疾病是由血液动力学紊乱驱动的。我决心把我的生命花在这项工作上。该奖项将提供关键的培训和发展时间，以确保我成功过渡到一个独立的教师职位。在Richard Hynes的实验室中，在我的F32奖下，我检查了纤连蛋白转录物对动脉内皮中低流量和干扰流量的调节，并发现了一个令人惊讶的剪接开关，导致包含两个广泛保守的替代外显子。纤维连接蛋白是动脉瘤中最丰富的细胞外基质蛋白之一。使用遗传模型，我表明，这种剪接开关保护动脉血管壁在低流量和干扰流量下的出血性剥离。尽管许多关键的内皮蛋白是选择性剪接的，并且选择性剪接程序正在成为其他发育过程中的关键控制机制，但对动脉内皮中选择性剪接的调节和功能几乎一无所知。为了解决这一缺陷，我将（目的1）更详细地研究纤连蛋白的选择性剪接保护免受解剖出血的机制，并（目的2）利用我们积累的数据库，在受干扰的流动下由巨噬细胞募集在内皮中诱导的选择性剪接事件，以确定关键的剪接因子及其在流动驱动的炎症中的功能。Richard Hynes（细胞外基质和纤连蛋白生物学专家）和Chris Burge（选择性剪接基因组规模分析专家）将在K99阶段指导我的工作。麻省理工学院拼接领域的人才聚集将是一个额外的好处;菲尔·夏普30年前在这里首次发现了拼接，它仍然是他的实验室和附近其他人的重点。这些目标的完成将为血流激活的动脉内皮的遗传调控增加新的维度，并通过对基础生物学的更深入理解，使血流驱动的血管疾病的诊断和治疗得以发展。

 DESCRIPTION (provided by applicant): Project Summary My long term goal is to lead a research group in a top tier academic institution in the pursuit of my interest: the genetic regulation of vascular development and disease, with a focus on the endothelium and hemodynamic activation. I am fascinated by the effects of blood flow on the vasculature and have focused on the transcriptional regulation of endothelial function for the past 10 years. I believe that in vivo genetics and imaging will be the key to understanding the complex interaction between the endothelium, blood flow, and recruited blood cells. This, in turn, is critical to understanding vascular development and diseases, such as aneurysms and atherosclerosis, which are driven by hemodynamic disturbance. I am committed to spending my life in this work. This award will provide critical training and development time to ensure my successful transition to an independent faculty position. In Richard Hynes' lab, under my F32 award, I examined the regulation of the Fibronectin transcript in response to low and disturbed flow in the arterial endothelium and uncovered a surprising splicing switch that resulted in the inclusion of two widely conserved alternative exons. Fibronectin is one of the most abundant extracellular matrix proteins in aneurysms. Using genetic models I showed that this splicing switch protects against hemorrhagic dissection of the arterial vessel wall under low and disturbed flow. Although a number of critical endothelial proteins are alternatively spliced, and alternative splicing programs are emerging as pivotal control mechanisms in other developmental processes, almost nothing is known about the regulation and function of alternative splicing in the arterial endothelium. To address this deficit, I will (Aim 1) examine i greater detail the mechanism(s) through which alternative splicing of Fibronectin protects against dissecting hemorrhage, and (Aim 2) exploit a database we have amassed, of alternative splicing events induced in the endothelium by macrophage recruitment under disturbed flow, to identify critical splicing factors and their functions in flow-­--driven inflammation. Richard Hynes (expert in extracellular matrix and Fibronectin biology) and Chris Burge (expert in genome-­--scale analysis of alternative splicing) will mentor my work during the K99 phase of the award. The congregation of talent in the splicing field at MIT will be an added benefit; Phil Sharp first discovered splicing here 30 years ago and it remains a focus of his lab and others nearby. The completion of these aims will add a new dimension to the genetic regulation of the flow-­--activated arterial endothelium, and enable the development of diagnostics and treatments for flow-­--driven vascular diseases through a deeper understanding of the basic biology.