Trans-Regulation of RNA-Binding Protein Motifs by MicroRNA

 描述（由申请人提供）：我们已经发现，微小RNA可以动态方式调节RNA结合蛋白（RBP）结合位点，并表明微小RNA以动态方式间接或直接调节RBP结合位点，从而增加转录后调控密码的复杂性和层次。这种模型，我们称之为“sxRNA”的结构相互作用的RNA，代表了一个令人兴奋的新的转录后调控范式，可能提供基因表达调控的基本方面。我们已经在信息上鉴定了几种“反式作用”sxRNA，我们的初步数据表明，阳性和阴性变异都是可能的。阴性或“阻遏物”sxRNA复合物的实例抑制RBP结合活性，而阳性或“诱导物”形式的sxRNA复合物可增加RBP结合，并且可使用RBP免疫沉淀（RIP）随后进行miRNA分析来捕获。这种新的调控机制可以解释如何复杂的基因多功能，包括选择性剪接的调控可以在转录后水平。此外，这些相互作用的组合可以以模块化和组合的方式使用，从而为转录后调控密码增加层次复杂性。与令人印象深刻的microRNA领域类似，sxRNA有可能增加我们对基因调控的理解，同时创造技术机会，利用这种调控作为组织特异性mRNA替代基因治疗，抗病毒，基于RNA的分子工具，诊断，甚至利用这些新发现的转录后方法的治疗。该提案的目标是测试一种新的转录后模型，其中miRNA和RBP竞争反式影响RNA结构。该项目旨在通过利用实验和信息学方法鉴定数十种（可能数百种）天然存在的microRNA-mRNA-RBP相互作用，表征sxRNA trans-RNA相互作用的结构性质及其对RBP结合的影响，并使用专门设计的MS 2-sxRNA报告基因验证它们在细胞系统中的存在和翻译作用。

 DESCRIPTION (provided by applicant): We have discovered that micoRNAs can modulate RNA-binding protein (RBP) binding sites in a dynamic manner and suggest that miRNAs indirectly or directly modulate RBP-binding sites in a dynamic manner adding complexity and hierarchy to a post-transcriptional regulatory code. This model, which we call "sxRNA" for structurally interacting RNA, represents an exciting new post-transcriptional regulatory paradigm that may provide fundamental aspects of gene expression regulation. We have informatically identified several "trans-acting" sxRNAs and our preliminary data suggests that both positive and negative variations are possible. Examples of negative or "repressor" sxRNA complexes inhibit RBP binding activity, while positive or "inducer" versions of sxRNA complexes may increase RBP binding and can be captured using RBP immunoprecipatation (RIP) followed by miRNA analysis. This novel regulatory mechanism may explain how complex regulation of gene multi-functionality including alternative splicing could be regulated at the post-transcriptional level. Furthermore, combinations of these interactions could be used in a modular and combinatorial manner adding a hierarchical complexity to the post-transcriptional regulatory code. Similar to the impressive microRNA field, sxRNA has the potential to increase our understanding of gene regulation while simultaneously creating technological opportunities to utilize this regulation as a tissue specific mRNA alternative to gene therapy, anti-viral, RNA-based molecular tool, diagnostic, or even therapeutic that exploits these newly discovered post-transcriptional approaches. The goal of this proposal is to test a new post-transcriptional model in which miRNA and RBPs compete in trans to influence RNA structure. This project is designed to identify dozens (possibly hundreds) of naturally occurring microRNA-mRNA-RBP interactions by exploiting both experimental and informatic approaches, characterize the structural nature of the sxRNA trans-RNA interaction and its effect on RBP binding and validate their existence and role in translation in a cellular system using a specifically designed MS2-sxRNA reporter.