Regulation of mRNA processing: Mechanisms and Consequences

mRNA 加工的调控：机制和后果

• 批准号: 10432005
• 负责人: James L. Manley
• 金额: $ 79.43万
• 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 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10432005
• 关键词: Affect; Amyotrophic Lateral Sclerosis; Anemia; Antiviral Response; Area; Brain; Complex; Coupled; Cytoplasm; Development; Disease; Event; GATA1 gene; Genes; Genetic Transcription; Goals; Homeostasis; Human; Lead; MAP3K7 gene; Malignant Neoplasms; Membrane Proteins; Messenger RNA; Motor Cortex; Mutation; NF-kappa B; Neoplasms; Neurodegenerative Disorders; Nuclear; Nuclear Inner Membrane; Nuclear Pore Complex; Nuclear Pore Complex Proteins; Open Reading Frames; Pathway interactions; Patients; Physiological; Polyadenylation; Pre-mRNA Polyadenylation Factor; Process; Protein Isoforms; Proteins; RNA; RNA Helicase; RNA Splicing; Reaction; Regulation; SARS-CoV-2 infection; Sampling; Specificity; Suggestion; Translating; Work; cell type; cofactor; design; exosome; experimental study; frontotemporal lobar dementia-amyotrophic lateral sclerosis; human embryonic stem cell; insight; mRNA Precursor; p38 Mitogen Activated Protein Kinase; sporadic amyotrophic lateral sclerosis

Synthesis of eukaryotic mRNAs is a complex process, and includes splicing and 3’ end formation of mRNA precursors. My lab has studied these processes for many years, including recently how they function in differentiation and disease. This proposal continues our studies on these topics, and can be divided into the following areas. mRNA processing in cancer. Work investigating how mutations in genes encoding splicing factors lead to MDS and other neoplasms will be continued, with an emphasis on SF3B1. Studies to elucidate both the mechanism(s) by which SF3B1 mutations affect splicing as well as the pathways that are dysregulated and lead to disease will be pursued. With respect to mechanism, an immediate goal will be to continue characterizing the SF3B1-interacting region of SUGP1. Another goal is to identify the SUGP1- interacting RNA helicase and determine its function in normal BP recognition, which will then allow determination of the detailed mechanism by which SF3B1 mutations disrupt splicing. With respect to pathways, experiments to elucidate the details of the recently described MAP3K7/p38/GATA1 pathway that underlies anemia in MDS will be performed. Missplicing events that affect other relevant pathways, such as aberrant activation of NF-kB, will be studied. FUS and other RBPs in ALS/FTD. With respect to FUS, experiments to analyze FUS nucleocytoplasmic homeostasis, a process important for formation of toxic cytoplasmic FUS aggregates, will be continued. Evidence supporting a gating mechanism involving the NPC and interactions with specific nucleoporins, themselves implicated in amyotrophic lateral sclerosis (ALS), will be pursued. Suggestions that this process may involve cell-type specificity, with MNs perhaps being more permissive, will be investigated. Experiments pursing the observation that RBP aggregates and consequent missplicing occur in a large fraction of sporadic ALS/FTD patient brains in the absence of known mutations will be continued. Aggregates will be isolated from motor cortex samples and protein/RNA composition determined to investigate what might nucleate their formation. Regulation of PA factor activity by AS. Experiments investigating the functions of isoforms of the polyadenylation (PA) factor WDR33 will be pursued. Two short isoforms, v2 and v3, are produced by intronic PA), and v2 but not v3 is an inner nuclear membrane protein not directly involved in PA. Interacting proteins of both will be identified, and results suggesting they are upregulated by an NF-kB pathway will be pursued. Evidence that the two isoforms function in the antiviral response, including to SARS-CoV-2 infection, will be further investigated. The finding that 45 isoforms of the PA factor Fip1 are produced in humans will be explored. eRNAs as mRNAs. Results suggesting that certain nuclear unstable lncRNAs, including eRNAs, are stabilized, exported to the cytoplasm and in some case translated, will be pursued. Observations that up to 5% of eRNAs contain significant ORFs and may be translated when the exosome cofactor Mtr4 is depleted will be further investigated. The possible physiological significance of eRNA “activation” will be studied in differentiated hESCs, which were shown to have sharply reduced levels of Mtr4.

真核mRNA的合成是一个复杂的过程，包括mRNA的剪接和3'端形成 前体我的实验室已经研究这些过程很多年了，包括最近它们在 分化与疾病这一建议继续我们对这些主题的研究，并可分为 以下地区。癌症中的mRNA加工。研究基因编码剪接的突变 将继续研究导致MDS和其他肿瘤的因素，重点是SF 3B 1。研究阐明 SF 3B 1突变影响剪接的机制以及 失调并导致疾病将被追求。在机制方面，一个直接目标是 继续表征SUGP 1的SF 3B 1相互作用区域。另一个目标是识别SUGP 1- 相互作用的RNA解旋酶，并确定其在正常BP识别中的功能，这将允许 确定SF 3B 1突变破坏剪接的详细机制。关于路径， 这些实验旨在阐明最近描述的MAP 3 K7/p38/GATA 1通路的细节， 将进行MDS贫血检查。影响其他相关途径的错误剪接事件，例如异常 将研究NF-kB的活化。ALS/FTD中的FUS和其他RBP。关于FUS， 分析FUS核质稳态，这是形成毒性细胞质FUS的重要过程 聚合，将继续。支持涉及NPC和相互作用的门控机制的证据 与肌萎缩侧索硬化症（ALS）有关的特定核孔蛋白的研究将继续进行。建议这一进程可能 涉及细胞类型特异性，MN可能更宽容，将被调查。实验 追求的观察，RBP聚集体和随之而来的错误剪接发生在一个大部分的散发性 在不存在已知突变的情况下，将继续ALS/FTD患者的脑研究。聚合物将与 运动皮层样本和蛋白质/RNA组成，确定调查什么可能使他们的细胞核化。 阵AS对PA因子活性的调节。实验研究的功能的异构体的 多聚腺苷酸化（PA）因子WDR 33。两个短的同种型，v2和v3，由内含子产生。 PA），v2而不是v3是不直接参与PA的内核膜蛋白。相互作用蛋白 两者都将被鉴定，并将追踪提示它们被NF-κ B途径上调的结果。 这两种亚型在抗病毒反应中发挥作用的证据，包括对SARS-CoV-2感染的证据， 进一步调查。PA因子Fip 1的45种亚型在人类中产生的发现将是 探讨了eRNA作为mRNA。结果表明，某些核不稳定的lncRNA，包括eRNA， 稳定化，输出到细胞质，并在某些情况下翻译，将继续进行。高达5%的观察结果 的eRNA含有显著的ORF，并且当外泌体辅因子Mtr 4被耗尽时可以被翻译。 进一步调查。eRNA“激活”的可能生理意义将在 分化的hESC，其显示具有急剧降低的Mtr 4水平。