Mechanisms of Transient Transcription in Yeast

酵母瞬时转录机制

• 批准号: 8135294
• 负责人: RANDY S STRICH
• 金额: $ 30.58万
• 依托单位: UNIV OF MED/DENT NJ-SCH OSTEOPATHIC MED
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8135294
• 关键词: Acetylation; Binding; Biochemical Genetics; Biological Assay; Cell division; Cells; Chromatin; Chromatin Remodeling Factor; Chromosome Segregation; Complex; Coupled; DNA Binding; DNA Damage; DNA Maintenance; DNA damage checkpoint; Deacetylation; Defect; Development; Embryo; Event; Failure; G2/M Transition; Gcn5p; Gene Expression; Genes; Genetic Transcription; Goals; Health; Histone Deacetylase; In Vitro; Lead; Ligase; Mediating; Meiosis; Meiotic Recombination; Mitotic; Modeling; Modification; Molecular; Monitor; Nucleosomes; Pathway interactions; Play; Proteins; Recruitment Activity; Regulation; Reporting; Repression; Role; SAGA; Saccharomycetales; Series; Signal Transduction; System; Testing; Time; Trans-Activators; Transcription Repressor/Corepressor; Transcriptional Activation; Yeasts; anaphase-promoting complex; gene induction; histone acetyltransferase; in vivo; prevent; programs; promoter; research study; response; transcription factor; ubiquitin ligase

DESCRIPTION (provided by applicant): The spatial and temporal expression of specific gene sets is critical for the execution of complex differentiation programs. The long-term goal of this study is to mechanistically define transient transcription in the context of the budding yeast meiotic differentiation program. Many genes required for the meiotic landmark events are repressed during mitotic cell division but then transiently induced during development in temporal waves termed "early", "middle" and "late". Ume6p binds "early" meiotic gene promoters and mediates their vegetative repression by recruiting both histone deacetylase (HDAC) and chromatin remodeling complexes. We have recently discovered that early meiotic gene induction requires Ume6p destruction by the Cdc20p- directed anaphase promoting complex/cyclosome (APC/C) ubiquitin ligase. Although well known for its role in controlling the G2 -> M transition in mitotic cells, this is the first report that APC/CCdc20 targets a transcription factor for destruction. In addition, Ume6p destruction is restricted to cells entering meiosis, even though APC/CCdc20 is active (and Ume6p is present) during mitotic cell division. The meiotic inducer Ime1p provides a trigger to promote Ume6p destruction thus suggesting a new mechanism by which APC/C substrate selection is redirected within the context of a differentiation program. While searching for potential meiosis-specific destruction signals, we discovered that Ume6p is a substrate of the Gcn5p histone acetyltransferase (HAT) complex called SAGA. The well-studied acetylation and deacetylation of nucleosomes maintains chromatin in "open" and "closed" configurations, respectively. However, the acetylation of transcription factors has not been demonstrated previously in yeast and has only been described for a few transactivators in mammalian systems. To our knowledge, Ume6p is the first transcriptional repressor found to be acetylated. Interestingly, preliminary results point to a role for acetylation in both preventing Ume6p DNA binding ability and enhancing its degradation. These findings suggest a new model for Gcn5p-dependent transcriptional activation through direct inhibition of Ume6p repressor function. Following induction, meiotic gene expression and the execution of landmark events are coupled by a series of checkpoint systems. Preliminary results indicate that Ume6p destruction is prevented upon activation of the DNA damage checkpoint. The mechanism by which Ume6p is protected from destruction following checkpoint activation, or the role of additional checkpoint pathways, is unknown. To understand the molecular mechanisms by which Ume6p-dependent repression is relieved upon meiotic induction, and how repression is reestablished in response to checkpoint pathways, the following aims are proposed: Aim1. Dissect the molecular mechanisms directing developmental re-tasking of the APC/C. Aim2. Determine the role that acetylation plays in Ume6p activity and regulation. Aim3. Identify and characterize the meiotic pathways that mediate Ume6p destruction. PUBLIC HEALTH RELEVANCE: During normal development, gene expression is precisely controlled to provide the correct gene product at the correct time. Failure to execute the proper gene expression program can lead to developmental defects or embryonic lethality. This proposal describes experiments to understand how these transcription programs are regulated when cells undergo development.

描述（由申请人提供）：特定基因组的空间和时间表达对于复杂分化程序的执行至关重要。这项研究的长期目标是在芽殖酵母减数分裂分化程序的背景下机械地定义瞬时转录。减数分裂里程碑事件所需的许多基因在有丝分裂细胞分裂期间受到抑制，但随后在称为“早期”、“中期”和“晚期”的时间波发育过程中短暂诱导。 Ume6p 结合“早期”减数分裂基因启动子，并通过招募组蛋白脱乙酰酶 (HDAC) 和染色质重塑复合物来介导其营养抑制。我们最近发现早期减数分裂基因诱导需要 Cdc20p 定向的后期促进复合物/环体 (APC/C) 泛素连接酶破坏 Ume6p。尽管 APC/CCdc20 以其在控制有丝分裂细胞 G2 -> M 转变中的作用而闻名，但这是 APC/CCdc20 以转录因子为目标进行破坏的第一份报告。此外，Ume6p 的破坏仅限于进入减数分裂的细胞，即使 APC/CCdc20 在有丝分裂细胞分裂过程中处于活跃状态（并且存在 Ume6p）。减数分裂诱导剂 Ime1p 提供了促进 Ume6p 破坏的触发器，从而提出了一种新机制，通过该机制在分化程序的背景下重定向 APC/C 底物选择。在寻找潜在的减数分裂特异性破坏信号时，我们发现 Ume6p 是称为 SAGA 的 Gcn5p 组蛋白乙酰转移酶 (HAT) 复合物的底物。经过充分研究的核小体乙酰化和脱乙酰化分别使染色质保持“开放”和“闭合”构型。然而，转录因子的乙酰化此前尚未在酵母中得到证实，并且仅在哺乳动物系统中描述了少数反式激活因子。据我们所知，Ume6p 是第一个被发现被乙酰化的转录抑制因子。有趣的是，初步结果表明乙酰化在阻止 Ume6p DNA 结合能力和增强其降解方面发挥着作用。这些发现提出了一种通过直接抑制 Ume6p 阻遏蛋白功能来实现 Gcn5p 依赖性转录激活的新模型。诱导后，减数分裂基因表达和标志性事件的执行通过一系列检查点系统耦合。初步结果表明，DNA 损伤检查点激活后，Ume6p 的破坏就被阻止了。检查点激活后 Ume6p 免受破坏的机制或其他检查点途径的作用尚不清楚。为了了解减数分裂诱导后 Ume6p 依赖性抑制得以缓解的分子机制，以及如何根据检查点途径重新建立抑制，提出以下目标：剖析指导 APC/C 发育重新分配的分子机制。目标2。确定乙酰化在 Ume6p 活性和调节中的作用。目标3。识别并表征介导 Ume6p 破坏的减数分裂途径。公共健康相关性：在正常发育过程中，基因表达受到精确控制，以在正确的时间提供正确的基因产物。未能执行正确的基因表达程序可能会导致发育缺陷或胚胎致死。该提案描述了了解细胞发育时这些转录程序如何受到调节的实验。