MICRORNA TARGETING IN HEART FAILURE

描述（由申请人提供）：microRNAs （miRs）是一种小的非编码rna，通过破坏和/或靶信使rna （mrna）的翻译抑制来调节蛋白质表达。与mrna一样，miRs的表达在心脏肥厚和心力衰竭中受到调节，可根据即时生理需求微调心肌细胞功能。单个miRs倾向于调节同一功能通路内的许多效应物，通过多个平行扰动产生一致的生理反应，因此是有吸引力的治疗靶点。然而，一种miR靶向数十种或数百种不同的mrna，以及在心脏病中miR和mrna的调节表达，使与心脏病相关的特定miR功能的描述复杂化。为了克服这一问题，我们开发了新的技术，将RNA测序在大规模并行的下一代平台上与心脏RNA诱导的信号复合物的体内miR编程相结合。此外，我们在30个人类miR的成熟miR序列中发现了常见的序列变异和罕见的突变，并证明种子序列和非种子序列miR变异都可以改变mRNA的靶向性。因此，我们假设miR对正常和病变心脏的影响是由表达miR的水平、表达mRNA的水平和由序列互补决定的miR-mRNA配对的效率决定的，miR序列的变化进一步改变了miR的作用，独立于miR和mRNA的表达水平。为了检验这些假设，我们首次提出了系统和全面地定义重要的miR-mRNA配对事件并确定其在不同病理生理条件下基因编程小鼠心脏中的功能后果的研究。我们将继续进行研究，以确定人类miR序列变异和突变对心脏miR- mrna靶向、靶蛋白表达和心脏结构和功能的影响。通过结合我们在人类基因组学，RNA测序以及小鼠遗传模型的生成和分析方面的专业知识，我们具有独特的优势来实现这些目标。我们的长期目标是将人类基因变异和心脏病的研究与体外细胞和体内小鼠实验系统相结合，以充分了解miRs对心脏的影响。

DESCRIPTION (provided by applicant): microRNAs (miRs) are small non-coding RNAs that regulate protein expression by destabilization and/or translational inhibition of target messenger RNAs (mRNAs). Like mRNAs, expression of miRs is regulated in cardiac hypertrophy and heart failure, fine-tuning cardiomyocyte function in response to immediate physiological demands. Single miRs tend to regulate numerous effectors within the same functional pathway, producing a coherent physiological response via multiple parallel perturbations, and are therefore attractive therapeutic targets. However, targeting of dozens or hundreds of different mRNAs by one miR, together with regulated expression of miRs and mRNAs in cardiac disease, complicates delineation of specific miR functions relevant to heart disease. We have developed new techniques combining RNA sequencing on massively parallel next generation platforms with in vivo miR programming of cardiac RNA-induced signaling complexes to overcome this problem. Additionally, we have identified common sequence variants and rare mutations within the mature miR sequence of 30 human miRs and demonstrated that both seed sequence and non-seed sequence miR variants can alter mRNA targeting. Accordingly, we hypothesize that miR effects on the normal and diseased heart are determined by the levels of expressed miRs, the levels of expressed mRNAs, and the efficiency of miR-mRNA pairing as determined by sequence complementarity, and that miR sequence variations further alter miR effects, independent of miR and mRNA expression levels. To examine these hypotheses we propose the first studies to systematically and comprehensively define important miR-mRNA pairing events and determine their functional consequences in genetically programmed mouse hearts under different pathophysiological conditions. We will follow with studies to determining the consequences of human miR sequence variants and mutations on cardiac miR-mRNA targeting, target protein expression, and cardiac structure and function. We are uniquely positioned to achieve these goals by combining our expertise in human genomics, RNA sequencing, and generation and analysis of murine genetic models. It is our long-term goal to integrate studies of human gene variation and cardiac disease with in vitro cell-based and in vivo murine experimental systems to fully understand the impact of miRs on the heart.