Quantitative Analysis of RNA Folding in Yeast

描述（由申请人提供）：RNA构象变化的动力学和平衡是在活细胞中发生的大量RNA介导反应的关键特征。在过去的二十年里，RNA酶或核糖酶为RNA结构-功能分析提供了黄金标准，因为它们通过催化活性揭示了它们的组装成功能结构。但是，尽管经过二十年的研究，我们对RNA组装的基本机制的理解仍然有限。发夹核酶编码在（-）TRSV卫星RNA中，是一个小的催化基序，催化序列特异性可逆自裂。发夹核酶的现有结构、生化和动力学数据，以及基于酵母的体内报告系统的可用性，使其成为研究细胞内RNA组装途径的理想模型。该系统将用于确定活细胞内转录和翻译过程中对核酶组装的动力学和热力学影响，并监测共转录核酶组装过程中rna是否在离散折叠区域内相互作用。最终，这项工作将提高我们对体内RNA组装基本原理的理解。

DESCRIPTION (provided by applicant): The dynamics and equilibria of RNA conformational changes are critical features of the multitude of RNA-mediated reactions that take place in living cells. RNA enzymes or ribozymes have provided the gold standard for RNA structure-function analyses for the past two decades because they reveal their assembly into functional structures through catalytic activity. But, despite two decades of research our understanding of the basic mechanisms of RNA assembly remains limited. The hairpin ribozyme, encoded in the (-)TRSV satellite RNA, is a small catalytic motif that catalyzes sequence-specific reversible self-cleavage. Existing structural, biochemical, and kinetic data for the hairpin ribozyme, and the availability of a yeast-based in vivo reporter system make this motif and ideal model for studying intracellular RNA assembly pathways. This system will be used to determine the kinetic and thermodynamic influences on ribozyme assembly during transcription and translation inside living cells, and to monitor whether RNAs interact within discrete folding domains during co-transcriptional ribozyme assembly. Ultimately this work will improve our understanding of the basic principles governing RNA assembly in vivo.