Structure and activity of a cellular IRES element

mRNA的翻译受到严格的调控，需要起始因子和核糖体的有序组装。 亚单位。某些病毒和细胞mRNA通过以下方式规避了翻译起始的传统要求： 利用被称为内部核糖体进入位点（IRES）的结构化RNA元件将核糖体引导到细胞内， 翻译起始位点。IRES结构与功能密切相关，但三维组织 对细胞的IRES了解甚少。该建议旨在获得IRES的晶体结构 Cyr61 mRNA中的元件与其功能所需的蛋白质复合，提供了第一个高分辨率 IRES单元的结构。Cyr 61是一种血管生成诱导剂，被认为在血管生成中起关键作用。 肿瘤进展。双顺反子报告基因检测将确定最小功能IRES和反式作用因子 将确定所需的活动。化学和酶的探测将决定次要的 Cyr61 IRES的结构，并确定适合结晶的结构元件。结果从这个 这项工作将产生对Cyr61 IRES的详细机制理解，并可能阐明 细胞IRES活性，可能确定治疗操纵Cyr61生产的策略。

Translation of mRNA is tightly regulated, requiring the ordered assembly of initiation factors and ribosomal subunits. Certain viral and cellular mRNAs circumvent traditional requirements for translation initiation by utilizing structured RNA elements termed internal ribosome entry sites (IRESs) to direct ribosomes to the translational start site. IRES structure is intimately linked to function, but the three-dimensional organization of cellular IRESs is poorly understood. This proposal aims to obtain a crystallographic structure of the IRES element in Cyr61 mRNA in complex with proteins required for its function, providing the first high-resolution structure of a cellular IRES element. Cyr61 is an angiogenic inducer suggested to play a critical role in tumor progression. Dicistronic reporter assays will define a minimal functional IRES and trans-acting factors required for activity will be identified. Chemical and enzymatic probing will determine the secondary structure of the Cyr61 IRES and identify structural elements amenable to crystallization. Results from this work will yield a detailed mechanistic understanding of the Cyr61 IRES and may elucidate mechanisms of cellular IRES activity, possibly identifying strategies to therapeutically manipulate Cyr61 production.