Regulation of mRNA processing: Mechanisms and consequences

mRNA 加工的调控：机制和后果

• 批准号: 9071558
• 负责人: James L. Manley
• 金额: $ 76.89万
• 依托单位: COLUMBIA UNIV NEW YORK MORNINGSIDE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-08 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9071558
• 关键词: Affect; Alternative Splicing; Area; C9ORF72; Code; Complex; Consensus; Coupled; DNA Damage; DNA Polymerase II; DNA-Binding Proteins; Defect; Development; Disease; Eukaryotic Cell; Event; G-Quartets; Gene Mutation; Genetic Transcription; Laboratories; Lead; Link; Malignant Neoplasms; Messenger RNA; Metabolism; Mutation; Neurodegenerative Disorders; Nuclear; Polyadenylation; Prevention; Process; Protein Isoforms; Proteins; RNA; RNA Helicase; RNA Processing; RNA Splicing; Reaction; Regulation; Role; Site; Structure; TCF3 gene; Transcript; Transcription Process; Work; design; disease-causing mutation; genetic regulatory protein; helicase; hnRNP-H; human embryonic stem cell; insight; leukemia; mRNA Precursor; overexpression; pluripotency; protein function; public health relevance; research study; stem; tumor

 DESCRIPTION (provided by applicant): Synthesis of mRNAs in eukaryotic cells is a highly complex process, including, in addition to transcription itself, splicing and 3' end formation of mRNA precursors. My laboratory has studied the mechanisms and regulation of these processes for many years, and also shown how they are integrated with other cellular events such as DNA damage, and also how they function in differentiation and disease. This proposal combines our studies in these areas, and can be divided into the following three broad areas: 1) mRNA processing and disease. Studies examining the roles of several proteins implicated in RNA metabolism in cancer and neurodegenerative disease will be pursued. The role of RNA processing factors in cancer reflects two distinct mechanisms, overexpression of splicing regulatory proteins and mutation of genes encoding various splicing factors. With respect to the former, ongoing experiments stemming from studies elucidating the mechanism underlying deregulation of PKM alternative splicing (AS) in cancer will be pursued. Important questions include the generality of the mechanism, the importance of splicing factor overexpression to tumor development, and the identity of the critical regulated splicing events. Splicing factor mutations can cause MDS and certain leukemias, and ongoing studies are aimed at elucidating mechanisms by which mutations affect the function of several, e.g., SRSF2 and SF3B1, and how such defects lead to disease. With respect to neurodegenerative disease, work centers on how ALS-causing mutations in the RNA/DNA-binding protein TLS/FUS disrupt protein function; how the hexanucleotide repeat expansions in C9ORF72 form G-quadruplex structures that sequester hnRNP H, disrupt AS, and contribute to ALS; and how disease-causing mutations in the RNA/DNA helicase Senataxin affect its role in prevention of transcription-induced DNA damage. 2) mRNA processing and differentiation. Studies examining how changes in AS and alternative polyadenylation (APA) contribute to human embryonic stem cell pluripotency and differentiation will be pursued. How AS affects the function of the transcriptional regulator TCF3 and how this contributes to differentiation will be determined. The mechanism of the AS event, including how the splicing regulators are themselves regulated, will be investigated. Our unexpected finding that AS of transcripts encoding two PA factors, both subunits of CPSF and implicated in AAUAAA recognition, is altered during differentiation will be pursued. How the distinct isoforms affect APA, and differentiation, will be determined, as will the mechanism by which they influence PA site choice. 3) RNA processing and transcription. Experiments examining the function of the RNA pol II CTD, a unique domain consisting of 26-52 heptad repeats (consensus YSPTSPS), will be continued. For example, recent experiments establishing a role for P-Tyr1 residues in facilitating turnover of certain lncRNAs, and suggesting that this involves interaction with the RNA helicase Mtr4 and the PA machinery, will be pursued. These and other experiments will advance our understanding of the "CTD code" that links transcription and RNA processing.

 描述（由适用提供）：真核细胞中mRNA的合成是一个高度复杂的过程，包括除了转录本身，剪接和mRNA前体的3'末端形成。多年来，我的实验室研究了这些过程的机制和调节，并展示了它们如何与其他细胞事件（例如DNA损伤）整合在一起，以及它们在分化和疾病中的作用。结合我们在这些领域的研究，可以分为以下三个广泛的领域：1）mRNA处理和疾病。将研究研究研究在RNA代谢中实施的几种蛋白质在癌症和神经退行性疾病中的作用的研究。 RNA加工因子在癌症中的作用反映了两种不同的机制，剪接调节蛋白的过表达以及编码各种剪接因子的基因突变。关于前者，将采用研究癌症中PKM替代剪接（AS）的机制的研究所引起的持续实验。重要的问题包括机制的一般性，剪接因子过表达对肿瘤发育的重要性以及严格调节的剪接事件的身份。剪接因子突变会导致MDS和某些白血病，并且持续的研究旨在阐明突变影响几种功能，例如SRSF2和SF3B1的功能，以及此类缺陷如何导致疾病。关于神经退行性疾病，工作集中在RNA/DNA结合蛋白TLS/FUS中的蛋白质蛋白功能中如何引起ALS的突变； C9orf72中的六核苷酸重复膨胀如何形成隔离HNRNP H的G四链体结构，破坏AS和对ALS的贡献；以及RNA/DNA解旋酶鼻毒素中引起疾病​​的突变如何影响其预防转录诱导的DNA损伤的作用。 2）mRNA处理和分化。研究如何进行AS AS和替代聚腺苷酸化（APA）的变化有助于人类胚胎干细胞多能和分化的研究。如何影响转录调节剂TCF3的功能以及如何确定这有助于分化的功能。将研究AS事件的机制，包括如何调节剪接调节器。我们出乎意料的发现，在编码两个PA因素的成绩单中，CPSF的亚基和AAUAAA识别中暗示的均应在分化过程中发生改变。不同的同工型如何影响APA和分化，以及它们影响PA位点选择的机制。 3）RNA处理和转录。将继续研究RNA POL II CTD功能的实验，这是一个由26-52个重复序列组成的独特域（共识YSPTSPS）。例如，最近的实验确定了P-TYR1的作用 这涉及与RNA解旋酶MTR4和PA机械的相互作用。这些和其他实验将提高我们对连接转录和RNA处理的“ CTD代码”的理解。