Mechanism for post-transcriptional gene regulation by Ribothrypsis

项目总结/摘要 基因表达的转录后调节是正常细胞稳态的基础。当前 生物学领域的一个模型是，mRNA主要作为“全长”分子存在， 翻译过程中核糖体的衰变。长期以来的理解是，mRNA衰变是由以下因素引发的： 去腺苷化，然后去帽，随后从两端进行核酸外切降解。共翻译 已知mRNA衰变主要发生在缺陷mRNA中。现有的下一代测序方法， mRNA降解谱靶向5'或3'末端，因此丢失了关于另一端的信息， 阐明了对mRNA衰变的新见解。在我的博士后培训期间，我开发了几个新的 转录组范围的测序方法，同时选择mRNA的两端。我发现了一本小说 涉及重复核酸内切裂解的典型mRNA的共翻译衰变机制 事件，由翻译核糖体介导，我们命名为“核糖体裂解”。Ribothrypsis是保守的 人类和酵母我们发现了意想不到的核糖体阶段的mRNA片段，并发现， 去腺苷化不是mRNA衰变的先决条件。我们对核糖体的发现表明， mRNA衰变比以前认为的更普遍。这一提议的核心假设是， 核糖体裂解是一种进化上保守的调节基因表达的机制， 许多因素来募集一种未知的核酸内切酶，我们称之为“核糖蛋白酶”。我们在此提议， 利用我们过去的发现，并利用尖端的新RNA测序方法，以实现 全面了解核糖体形成及其调控的机制基础。在这 根据这一提议，我们将研究（i）诱导核糖体停滞的细胞条件对RNA衰变的影响 中间体;（ii）其他真核生物中核糖胞壁形成的保守性;（iii）核糖胞壁形成蛋白的身份;和（iv） RNA修饰在触发核糖体裂解中的作用。这些目标反映在我们的长期目标中， 了解人类疾病中RNA衰变失调的机制。从分子角度 这一提议的获得也可以拓宽我们对核糖体生物学和RNA修饰的理解。

PROJECT SUMMARY/ ABSTRACT Post-transcriptional regulation of gene expression is fundamental to normal cellular homeostasis. The current model in the field of biology is that mRNAs exist primarily as ‘full-length’ molecules that are ‘protected’ from decay by ribosomes during translation. The long-established understanding is that mRNA decay is initiated by deadenylation followed by decapping and subsequent exonucleolytic decay from both ends. Co-translational mRNA decay is known to occur mainly in defective mRNAs. Existing next-generation sequencing methods that profile mRNA decay target either the 5’ or 3’ ends, thus missing information about the other end that could illuminate new insights into mRNA decay. During my post-doctoral training, I developed several novel transcriptome-wide sequencing methods that concurrently select both ends of mRNAs. I discovered a novel mechanism of co-translational decay of canonical mRNAs that involves repeated endonucleolytic cleavage events, mediated by translating ribosomes that we named “ribothrypsis”. Ribothrypsis is conserved between humans and yeast. We identified the unexpected ribosome-phased mRNAs fragmentation and found that deadenylation is not a prerequisite for mRNA decay. Our discovery of ribothrypsis revealed that co-translational mRNA decay is more widespread than previously thought. The central hypothesis of this proposal is that ribothrypsis is an evolutionarily conserved mechanism for modulating gene expression that can be triggered by numerous factors to recruit an unknown endonuclease that we termed “ribothrypsin”. We propose here to capitalize on our past discoveries and leverage cutting-edge novel RNA sequencing methods to achieve a comprehensive understanding of the mechanistic underpinnings of ribothrypsis and its regulation. In this proposal, we will investigate (i) the impact of cellular conditions that induce ribosome stalling on RNA decay intermediates; (ii) the conservation of ribothrypsis in other eukaryotes; (iii) the identity of ribothrypsin; and (iv) the role of RNA modifications in triggering ribothrypsis. These goals are mirrored by our long-term objective to understand the mechanisms that underlie RNA decay dysregulation in human diseases. The molecular insights gained in this proposal could also broaden our understanding of ribosome biology and RNA modifications.