Beta-arrestin signaling and microRNA biogenesis in cardioprotection

描述（由申请人提供）：心力衰竭（HF）是影响500多万美国人的主要公共卫生问题。心衰的总医疗费用每年超过300亿美元，预计随着人口老龄化还会增加。因此，迫切需要制定预防和治疗心衰的创新策略。- 1-肾上腺素能受体（- 1AR，一种7跨膜G蛋白偶联受体）信号传导对心功能的交感调节至关重要，但在心衰进展过程中，慢性儿茶酚胺刺激会对心功能产生有害影响。最近，人们认识到- 1AR信号通路涉及多种途径，包括一种新的心脏保护信号（CPS）途径，其中- 1AR使用-抑制蛋白（-arr）在缺乏G蛋白激活的情况下促进心肌细胞（CM）的存活。然而，下游核过程中α -arr介导的α - 1AR信号赋予心脏保护的分子机制尚不清楚。基于越来越多的人认识到microRNAs （miRs）是心血管功能的重要调节因子，我们假设miR可能代表了被低估的调节-arr介导的心脏保护的下游机制。我们的初步数据显示，1ar介导的arr1激活增强了miR的加工和几种已知调节细胞存活和凋亡途径的miR的表达。然而，增加-arr1调节的miRs （- 1-miRs）对心功能和组织损伤结果的重要性尚不清楚，也是当前研究的目标。本应用的目的是确定1-miR， miR-150及其两个CM靶基因（促凋亡egr2和p2x7r）在1- ar介导的1 -arrestin1 CPS中的重要性。核心假设是，-arr1介导的- 1AR调节机制通过miR-150加工和两个靶基因表达的改变，赋予心肌梗死（MI）和慢性儿茶酚胺刺激的心脏保护作用。在大量初步数据的指导下，我们的假设将通过追求三个具体目标来验证：1)确定miR-150是否调节冠状动脉结扎和慢性儿茶酚胺刺激的心功能反应。对心肌梗死和慢性儿茶酚胺治疗的体内功能丧失和功能获得方法将在小鼠中使用。2)确定miR-150的表达增加是否有助于卡维地洛（一种arr偏倚的阻滞剂）介导的心脏保护。遗传和药理学方法将用于CM和全心。3)阐明miR- 150功能性CM靶点egr2和p2x7r在HF中的作用。靶点的功能丧失和功能获得研究将显示它们在CM生存和心脏保护中的重要性。这项工作是创新的，因为从未研究过-arr信号对miR表达的调节。我们将通过“-arrestin1偏倚”的1AR信号通路发现miR加工和功能的新调控机制，并确定参与CPS的潜在重要miR靶标对。这项研究具有重要意义，因为1AR对心肌保护miR/促凋亡靶标对的调控能力为心衰的药理学操作带来了希望。最终，这些知识有可能为预防和治疗各种心脏疾病的新方法的发展提供信息。

DESCRIPTION (provided by applicant): Heart failure (HF) is a major public health problem affecting more than 5 million Americans. The total healthcare costs of HF are over $30 billion annually, and are projected to increase as the population ages. Therefore, there is a critical need to develop innovative strategies for prevention and treatment of HF. �1-adrenergic receptor (�1AR, a seven transmembrane G protein-coupled receptor) signaling is critical to sympathetic regulation of cardiac function, but becomes deleterious in response to chronic catecholamine stimulation during the progression of HF. More recently, it became appreciated that �1AR signaling involves multiple pathways including a novel cardioprotective signaling (CPS) pathway, in which �1AR uses �-arrestin (�-arr) to promote cardiomyocyte (CM) survival in the absence of G protein activation. However, the molecular mechanisms in downstream nuclear processes by which �-arr-mediated �1AR signaling confers cardiac protection are not well understood. Based on increasing recognition of microRNAs (miRs) as important regulators of cardiovascular function, we postulated that miR may represent under-appreciated downstream mechanisms regulating the �-arr- mediated cardioprotection. Our preliminary data show that �1AR-mediated activation of �-arr1 enhanced miR processing and expression of several miRs that are known to regulate cell survival and apoptotic pathways. However, the importance of increased �-arr1-regulated miRs (�1-miRs) to the outcome of cardiac function and tissue injury is unknown and a goal of the current study. The objective of this application is to define the importance of a �1-miR, miR-150 and two of its CM target genes (pro-apoptotic egr2 and p2x7r) in �1AR-mediated �-arrestin1 CPS. The central hypothesis is that �-arr1- mediated �1AR regulatory mechanisms confer cardioprotection against myocardial infarction (MI) and chronic catecholamine stimulation via miR-150 processing and altered expression of the two target genes. Guided by extensive preliminary data, our hypothesis will be tested by pursuing three specific aims: 1) Determine if miR-150 modulates cardiac functional responses to coronary ligation and chronic catecholamine stimulation. In vivo loss- and gain-of-function approaches in response to MI and chronic catecholamine treatment will be employed in mice. 2) Determine if increased expression of miR-150 contributes to carvedilol (a �-arr-biased �-blocker)-mediated cardioprotection. Both genetic and pharmacological approaches will be used in CM and whole hearts. 3) Elucidate the roles of egr2 and p2x7r, functional CM targets of miR- 150, in HF. Loss- and gain-of-function studies of targets will be performed to show their importance in CM survival and cardiac protection. The proposed work is innovative because the regulation of miR expression by �-arr signaling has never been studied. We will discover novel regulatory mechanisms of miR processing and function by �-arrestin1-biased �1AR signaling pathways and identify potentially important miR-target pairs that are involved in CPS. The proposed re- search is significant because the ability of �-arr1-biased agonism of �1AR to regulate cardioprotective miR/pro-apoptotic target pairs holds promise for pharmacological manipulation of HF. Ultimately, such knowledge has the potential to inform the development of novel approaches for the prevention and treatment of a variety of cardiac diseases.