Context-specific Functions of CDK8

CDK8 的上下文特定功能

• 批准号: 10549761
• 负责人: Jun-yuan Ji
• 金额: $ 33.9万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10549761
• 关键词: Abbreviations; Binding Proteins; Biochemical; Biological; Biology; Cardiovascular Diseases; Cell Cycle; Cell Line; Child; Clinical; Colorectal Cancer; Complex; Coupled; Cyclin-Dependent Kinases; DNA Binding; Data; Development; Disease; Drosophila genus; Drosophila melanogaster; E2F transcription factors; EGF gene; Epidermal Growth Factor; Epidermal Growth Factor Receptor; Eukaryota; Factor Analysis; Funding; G1 Phase; G1/S Transition; Gene Expression; Genes; Genetic; Genetic Screening; Genetic Transcription; Genome Stability; Genomics; Goals; Homeostasis; Human; Immunoprecipitation; Intake; Length; Link; Malignant Neoplasms; Mammals; Mass Spectrum Analysis; Mediator; Modeling; Mutate; Nutrient; Pathologic; Pathway interactions; Phase Transition; Phenocopy; Phenotype; Phosphotransferases; Polymerase; Positioning Attribute; Protein Analysis; Proteins; Proteomics; RNA Polymerase II; Receptor Signaling; Regulation; Reporting; Repression; Research; Response Elements; Retrotransposon; Role; Serum Response Factor; Signal Pathway; Sterols; System; Telomere Capping; Testing; Trans-Activators; Transforming Growth Factor beta; Up-Regulation; Wing; cofactor; cyclin C; design; ecdysone receptor; fly; gene repression; genetic analysis; genomic locus; human disease; in vivo; inhibitor; lipid biosynthesis; lipid metabolism; melanoma; model organism; mutant; novel; public health relevance; receptor; restraint; synergism; telomere; transcription factor; transcriptome sequencing

Title: Context-specific functions of CDK8 Project Summary: The long-term goal of this project is to elucidate the function and regulation of the CDK8 module, a key component of the transcription cofactor Mediator complex, in the versatile model organism Drosophila melanogaster. The four subunits of the CDK8 module – CDK8, CycC, MED12, and MED13 – are either mutated or amplified in cardiovascular diseases and a number of human cancers, such as melanoma and colorectal cancers. Elucidating the function and regulation of the CDK8 module in different biological contexts is essential to understanding the pathological consequences of CDK8 module misregulation, which is important for the design of clinical strategies to treat these diseases. Studies in the previous funding cycle of this project have demonstrated that CDK8-CycC serves as a critical regulatory node linking nutrient intake to fat metabolism and developmental timing in Drosophila development. These studies have shown that CDK8-CycC is a direct inhibitor of SREBP (sterol response element binding protein)-dependent gene expression, and that CDK8-CycC positively regulates ecdysone receptor-activated gene expression. The overall objective of this project is to unravel the function and regulation of CDK8 in different developmental contexts. Studies in the previous funding cycle illustrate that Drosophila is an ideal and powerful experimental system to achieve this long-term goal. Two new aims are proposed in this funding cycle. Aim 1 will determine the role of CDK8 in regulating the expression of telomeric retrotransposons and telomere biology in Drosophila. Our RNA-seq analyses revealed a specific upregulation of telomeric retrotransposon expression and a significant increase in telomere length in cdk8 and cycC mutants. We have also discovered similar deregulation of telomeric retrotransposon expression and telomere length in med7 and Scalloped (Sd) mutants. The transcription factor Sd functions downstream of the conserved Hippo pathway. Thus we propose to examine the unexplored functions of the CDK8 module and Sd in regulating the expression of telomeric retrotransposons and telomere length in Drosophila. Aim 2 of the proposal will identify and validate upstream regulators and downstream effectors of CDK8 in Drosophila. We have established novel and robust wing phenotypes caused by specific alterations of CDK8 activities, allowing us to perform a dominant modifier genetic screen to identify factors interacting with CDK8 in vivo. We have identified 26 genomic loci whose haploinsufficiency modifies these CDK8-specific phenotypes; further genetic analysis led us to identify genetic interactions between CDK8 and the components of the epidermal growth factor receptor (EGFR) and Dpp/TGFβ signaling pathways, as well as several specific genes. In parallel, we have performed immunoprecipitation coupled to mass spectrometry (IP- MS) analyses to identify proteins that can interact with CDK8. Combination of these biochemical and genetic screens puts us in a unique position to systematically identify and validate upstream regulators and down- stream effectors of CDK8 in vivo, which will impact our understanding of the context-specific functions of CDK8.

标题：CDK8的上下文特定功能 项目摘要： 该项目的长期目标是阐明CDK8模块的功能和调节，一个密钥 在多功能模型有机体果蝇中，转录辅因子介质复合物的组成部分 Melanogaster。 CDK8模块的四个亚基 - CDK8，CYCC，MED12和MED13 - 是 在心血管疾病和许多人类癌症（例如黑色素瘤）中突变或扩增 结直肠癌。在不同的生物学环境中阐明CDK8模块的功能和调节 对于理解CDK8模块不调节的病理后果至关重要，这很重要 为了设计治疗这些疾病的临床策略。该项目以前的资金周期的研究 已经证明CDK8-CYCC是将营养摄入与脂肪联系起来的关键调节节点 果蝇发展中的代谢和发展时机。这些研究表明CDK8-CYCC 是SREBP（固醇反应元件结合蛋白）依赖性基因表达的直接抑制剂，并且 CDK8-CYCC积极调节ecdysone受体激活的基因表达。总体目标 项目是在不同的发育环境中阐明CDK8的功能和调节。研究 以前的资金周期说明果蝇是一个理想且强大的实验系统 长期目标。在这个融资周期中提出了两个新目标。 AIM 1将确定CDK8在 调节果蝇中远程逆转录子和端粒生物学的表达。我们的RNA-Seq 分析表明，远程循环转座子的表达的特定上调和显着增加 CDK8和CYCC突变体中的端粒长度。我们还发现了远程仪的类似管制 Med7和扇贝（SD）突变体中的逆转录座子表达和端粒长度。转录因子 配置的河马途径下游的SD功能。我们建议检查意外 CDK8模块和SD的功能在调节中的表达 果蝇的长度。该提案的目标2将识别和验证上游监管机构和下游 果蝇中CDK8的效应子。我们已经建立了由特定的新颖而健壮的机翼表型 CDK8活动的改变，使我们能够执行主导的修饰遗传筛选以识别因素 与体内CDK8相互作用。我们已经确定了26个基因组基因座，其单倍弥补可修改这些 CDK8特异性表型；进一步的遗传分析使我们确定了CDK8和 表皮生长因子受体（EGFR）和DPP/TGFβ信号通路的成分以及 几个特定基因。同时，我们进行了与质谱法相连的免疫沉淀（IP- MS）分析以识别可以与CDK8相互作用的蛋白质。这些生化和遗传的结合 屏幕使我们处于独特的位置，可以系统地识别和验证上游监管机构和下游 - CDK8在体内的流效应，这将影响我们对CDK8特定于上下文功能的理解。