Mechanism of yeast gene regulation

酵母基因调控机制

• 批准号: 10646455
• 负责人: KEVIN STRUHL
• 金额: $ 81.02万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10646455
• 关键词: 3' Untranslated Regions; Address; Affect; Area; Binding; Binding Proteins; Biochemical; Biological; Biological Phenomena; Biological Process; Chemicals; Chromatin Loop; Complex; DNA Sequence; Development; Disease; Elements; Eukaryota; Evolution; Experimental Genetics; Gene Expression; Gene Expression Regulation; Genetic Transcription; Goals; Histone Acetylation; Histones; Human; Human Biology; Individual; Knowledge; Length; Measurement; Measures; Mediating; Messenger RNA; Methodology; Molecular; Molecular Genetics; Mutation; Nature; Noise; Organism; Pattern; Physiological; Poly A; Polyadenylation; Prevention; Process; Protein Isoforms; Proteins; RNA; Regulation; Ribosomal Proteins; Role; Structure; TAF1 gene; Transcription Coactivator; Transcription Initiation; Transcription Process; Transcriptional Regulation; Variant; Work; Yeasts; cell growth; functional genomics; human disease; improved; mRNA Stability; novel; promoter; protein protein interaction; recruit; response; transcription factor

PROJECT SUMMARY The mechanisms by which eukaryotes regulate gene expression are important for understanding many complex biological phenomena including human diseases. Prevention and treatment of such diseases have been and will continue to be improved by basic knowledge of gene regulation, especially because molecular mechanisms of transcriptional initiation are highly conserved in eukaryotic organisms ranging from human to yeast. This proposal will continue to investigate basic issues concerning molecular mechanisms of transcriptional regulation, polyadenylation, and mRNA stability in yeast, by combining molecular genetic, biochemical, functional genomic, and evolutionary approaches. Work in the first two sections will take advantage of our novel and recently developed methodologies for measuring half-lives, structure (DREADS via chemical probes), protein binding (CLIP-READS), and poly(A) length (A-READS) of individual mRNA 3' isoforms. First, in the area of polyadenylation, we will A) address the mechanisms for why polyadenylation is restricted to the 3' UTR. B) identify factors that are responsible for the wild-type poly(A) pattern, C) determine the factors and mechanistic basis for regulated polyadenylation during the diauxic shift (and perhaps other conditions), and D) elucidate 3'-isoform variation and regulation of poly(A) length. Second, for studies of mRNA stabilization/destabilization elements and half-lives of 3' isoforms and, we will A) perform RNA structural analysis during the degradation process, B) identify protein factors mediating the large differences in mRNA isoform stabilities C) perform directed genetic experiments to address how secondary structure affects mRNA stability, D) identify mRNA stabilization and destabilizing elements that differentially affected by environmental conditions, and E) identify factors important for regulated mRNA half-lives, which the goal of elucidating the mechanism of regulated mRNA stability. Third, we will address a variety of issues concerning transcriptional regulation including A) the nature of the transcriptional activator that coordinately regulates ribosomal protein gene expression via recruitment of TFIID, B) determining the mechanistic basis of why activator proteins do not function when bound downstream of or far away from the core promoter, C) DNA looping mechanisms, particularly the nature of the protein-protein interactions needed to form the loop and to stimulate transcription, and D) examining the role of histone acetylation in transcriptional regulation by generating non-acetylable derivatives of the 4 histones. Fourth, we will use a novel conceptual and experimental approach to distinguish biological function from biological noise that is based on a comparison of physiological responses, RNA and transcription factor binding profiles, and effects of mutations in yeast species of varying evolutionary distance. We will explicitly measure biological noise by making functional measurements of evolutionary irrelevant or random-sequence DNA in yeast. Overall, the proposal will answer fundamental questions about the interlinked processes of transcription, polyadenylation, and mRNA stability in a mechanistic and evolutionary framework.

项目摘要 真核生物调节基因表达的机制对于了解许多生物学过程非常重要。 包括人类疾病在内的复杂生物现象。这些疾病的预防和治疗 基因调控的基础知识已经并将继续得到改善，特别是因为分子 转录起始机制在从人类到 酵母本建议将继续研究有关分子机制的基本问题， 酵母中的转录调节、多聚腺苷酸化和mRNA稳定性，通过结合分子遗传学， 生物化学、功能基因组学和进化方法。前两部分的工作将需要 我们的新的和最近开发的方法的优势，用于测量半衰期，结构（DREADS通过 化学探针）、蛋白结合（CLIP-READS）和单个mRNA 3'端的多聚腺苷酸长度（A-READS） 同种型。首先，在多聚腺苷酸化领域，我们将A）解决为什么多聚腺苷酸化是 限制于3' UTR。B）鉴定导致野生型poly（A）模式的因子，C）确定 在二倍体转换过程中调节多聚腺苷酸化的因素和机制基础（也许还有其他因素）， 条件），和D）阐明3 '-同种型变异和多聚腺苷酸长度的调节。第二，研究 mRNA稳定化/不稳定化元件和3'亚型的半衰期，我们将A）进行RNA结构分析， 分析降解过程中，B）鉴定介导mRNA大差异的蛋白质因子 C）进行定向遗传实验以解决二级结构如何影响mRNA 稳定性，D）鉴定受环境差异影响的mRNA稳定化和去稳定化元件 E）鉴定对于调节的mRNA半衰期重要的因素，这是阐明mRNA半衰期的目的。 调节mRNA稳定性的机制。第三，我们将讨论有关转录的各种问题， 调节包括A）协同调节核糖体蛋白的转录激活因子的性质 通过募集TFIID的基因表达，B）确定激活蛋白为什么不 C）DNA成环机制， 特别是形成环和刺激转录所需的蛋白质-蛋白质相互作用的性质， 和D）通过产生不可乙酰化的组蛋白， 4组蛋白的衍生物。第四，我们将使用一种新颖的概念和实验方法来区分 生物噪声的生物学功能是基于生理反应，RNA和 转录因子结合谱，以及不同进化距离的酵母物种中突变的影响。 我们将明确地测量生物噪音，通过对进化无关或 酵母中的随机序列DNA。总的来说，该提案将回答有关相互关联的 转录，多聚腺苷酸化和mRNA稳定性的机制和进化框架的过程。

项目成果

How is polyadenylation restricted to 3'-untranslated regions?

聚腺苷酸化如何限制在 3-非翻译区域？

• DOI: 10.1002/yea.3915
• 发表时间: 2024
• 期刊: Yeast (Chichester, England)
• 作者: Struhl,Kevin

Condition-specific 3' mRNA isoform half-lives and stability elements in yeast.

• DOI: 10.1073/pnas.2301117120
• 发表时间: 2023-05-02
• 期刊: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• 影响因子: 11.1
• 作者: V. Geisberg, Joseph;Moqtaderi, Zarmik;Struhl, Kevin
• 通讯作者: Struhl, Kevin

Comparison of transcriptional initiation by RNA polymerase II across eukaryotic species.

• DOI: 10.7554/elife.67964
• 发表时间: 2021-09-13
• 期刊: eLife
• 影响因子: 7.7
• 作者: Petrenko N;Struhl K
• 通讯作者: Struhl K

The transcriptional elongation rate regulates alternative polyadenylation in yeast.

• DOI: 10.7554/elife.59810
• 发表时间: 2020-08-26
• 期刊: eLife
• 影响因子: 7.7
• 作者: Geisberg JV;Moqtaderi Z;Struhl K
• 通讯作者: Struhl K

A compensatory link between cleavage/polyadenylation and mRNA turnover regulates steady-state mRNA levels in yeast.

• DOI: 10.1073/pnas.2121488119
• 发表时间: 2022-01-25
• 期刊: Proceedings of the National Academy of Sciences of the United States of America
• 影响因子: 11.1
• 作者: Moqtaderi Z;Geisberg JV;Struhl K
• 通讯作者: Struhl K