MICRORNA TARGETING IN HEART FAILURE

描述（由申请人提供）：微小RNA（miR）是通过靶信使RNA（mRNA）的去稳定化和/或翻译抑制来调节蛋白质表达的小的非编码RNA。与mRNA一样，miR的表达在心脏肥大和心力衰竭中受到调节，微调心肌细胞功能以响应即时的生理需求。单个miR倾向于调节相同功能途径内的众多效应物，通过多个平行扰动产生连贯的生理反应，因此是有吸引力的治疗靶标。然而，一种miR靶向数十种或数百种不同的mRNA，以及心脏疾病中miR和mRNA的调节表达，使与心脏疾病相关的特定miR功能的描述复杂化。我们已经开发了新技术，将大规模并行下一代平台上的RNA测序与心脏RNA诱导的信号传导复合物的体内miR编程相结合，以克服这个问题。此外，我们已经在30种人类miR的成熟miR序列中鉴定了常见的序列变体和罕见的突变，并证明种子序列和非种子序列miR变体都可以改变mRNA靶向。因此，我们假设miR对正常和患病心脏的作用是由表达的miR水平、表达的mRNA水平和由序列互补性确定的miR-mRNA配对效率决定的，并且miR序列变异进一步改变miR作用，而与miR和mRNA表达水平无关。为了检验这些假设，我们提出了第一个研究，系统和全面地定义重要的miR-mRNA配对事件，并确定其功能的后果，在不同的病理生理条件下的遗传编程小鼠心脏。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，靶蛋白表达以及心脏结构和功能的影响。通过结合我们在人类基因组学、RNA测序以及鼠遗传模型生成和分析方面的专业知识，我们处于独特的地位，能够实现这些目标。我们的长期目标是将人类基因变异和心脏疾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miR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.