The Positive Roles of Lysine Deacetylase Complexes in Regulating Transcriptional Dynamics (Diversity Supplement)

项目总结 而传统的转录模型将赖氨酸乙酰转移酶(KATS，也称为HATS)转换为反式-乙酰基转移酶。 作为辅抑制子的书面辅活化子和赖氨酸脱乙酰酶(KDAC，也称为HDAC)，是一种- 大量证据表明，KDAC可以以基因依赖的方式促进转录。然而， 其转录促进功能的潜在机制还知之甚少。的长期目标是 我们的工作是确定KDAC、KATS和乙酰化调节信号调节的转导的机制。 脚本。这项拟议的研究利用糖皮质激素信号作为模型系统。该实验室发表的研究成果 证明KDAC不仅是糖皮质激素介导的转录抑制所必需的，也是 靶基因的转录激活。初步研究表明，GR激活需要KDAC1 根据靶基因的不同，通过多种机制转录，作用于 RNA聚合酶II(RNA POL II)募集到转录起始点(TSS)。下一个合乎逻辑的步骤是确定 KDAC1复合体参与并研究它们在糖皮质激素受体(GR)激活转录中的作用。 母基金(R01GM139829)的目标是定义KDAC在运输动态中的角色。 糖皮质激素受体(GR)激活基因的转录循环。所描述的研究计划的目标 在这个多样性补充中是为了研究KDAC活性对RNA聚合酶II动力学的影响 与GR激活基因的相关性。这一假设将通过三个具体目标进行实验验证。 在第一个特定目标中，I类KDAC对GR激活的转录爆发动力学的影响 将测量基因，以确定受KDAC活性上游或下游调节的GR激活基因。 转录起始流，基于KDAC促进的机制的假设 转录是基因特有的。在第二个特定目的中，KDAC活性对RNAPolII分布的影响 GR激活基因的增强子、启动子和基因体上的新生转录模式将 在全基因组范围内被确定。工作假说是KDAC活性调节转录和RNA Poll II以基因特有的方式在不同基因区域的动态。在第三个特定目标中，组蛋白的动力学 乙酰化和rna polii将在GR激活的基因上进行研究，在该基因上可以看到转录。 在活细胞中。这将使用纳米抗体来检测组蛋白乙酰化和活细胞中的RNA PolII。 这项工作将产生有关KDAC转录功能的新的机械性知识 对细胞过程的基本了解以及通过调节表观来治疗疾病- 诺姆。重要的是，在进行这项研究的过程中，研究生将得到 补充剂将获得对在生物医学劳动力中成功职业生涯至关重要的技能。

PROJECT SUMMARY Whereas traditional models of transcription cast lysine acetyltransferases (KATs, also known as HATs) as tran- scriptional coactivators and lysine deacetylases (KDACs, also known as HDACs) as corepressors, an abun- dance of evidence demonstrates that KDACs can facilitate transcription in a gene-dependent fashion. However, the mechanisms underlying their transcription-promoting functions are poorly understood. The long-term goal of our work is to define mechanisms by which KDACs, KATs, and acetylation regulate signaling-modulated tran- scription. The proposed study utilizes glucocorticoid signaling as a model system. Published studies from the lab demonstrate that KDACs are required not only for glucocorticoid-mediated transcriptional repression but also for transcriptional activation of target genes. Preliminary studies show that KDAC1 is required for GR-activated transcription by multiple mechanisms, depending on the target gene, acting either upstream or downstream of RNA polymerase II (RNA Pol II) recruitment to the transcription start site (TSS). The next logical step is to identify the KDAC1 complexes involved and investigate their roles in glucocorticoid receptor (GR)-activated transcription. The objective of the parent grant (R01GM139829) is to define the roles of KDACs in the dynamics of the tran- scriptional cycle at glucocorticoid receptor (GR)-activated genes. The objective of the research plan described in this diversity supplement is to investigate the impact of KDAC activity on the dynamics of RNA polymerase II association with GR-activated genes. This hypothesis will be tested experimentally through three specific aims. In the first specific aim, the impact of Class I KDACs on the kinetics of transcriptional bursting at GR-activated genes will be measured to identify GR-activated genes that are regulated by KDAC activity upstream or down- stream of transcriptional initiation, working from the hypothesis that the mechanism by which KDACs facilitate transcription is gene-specific. In the second specific aim, the impact of KDAC activity on RNA pol II distribution and the pattern of nascent transcription at enhancers, promoters, and gene bodies of GR-activated genes will be determined genome-wide. The working hypothesis is that KDAC activity regulates transcription and RNA pol II dynamics at distinct gene regions in a gene-specific fashion. In the third specific aim the dynamics of histone acetylation and RNA pol II will be investigated at a GR-activated gene at which transcription can be visualized in live cells. This will be performed using nanobodies to detect histone acetylation and RNA pol II in live cells. This work will generate novel mechanistic knowledge of the transcriptional functions of KDACs that is relevant to basic understanding of cellular processes as well as treatment of disease through modulation of the epige- nome. Importantly, in the process of doing this research the graduate student who will be supported by the supplement will gain skills critical to a successful career in the biomedical workforce.