Mechanism of Transcription Control by Drosophila TAF250

果蝇TAF250的转录控制机制

• 批准号: 6530212
• 负责人: DAVID WASSARMAN
• 金额: $ 28.54万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6530212
• 关键词: DNA binding protein; DNA directed RNA polymerase; Drosophilidae; binding sites; biological signal transduction; biotechnology; chromatin; gene expression; gene mutation; genetic library; genetic promoter element; genetic screening; genetic transcription; genetically modified animals; microarray technology; molecular site; phosphorylation; polymerase chain reaction; protein isoforms; protein protein interaction; protein structure function; radionuclide double label; temperature sensitive mutant; transcription factor; yeast two hybrid system

DESCRIPTION (provided by applicant): TAFII250 is one of more than 10 TATA-binding protein (TBP)-associated factors (TAFIIs) that are complexed with TBP in TFIID. Binding of TFIID to the core promoter surrounding the transcription start site of a gene nucleates assembly of the preinitiation complex (PlC), which contains RNA polymerase II (Pol II) and the general transcription factors TFIIA, TFIIB, TFIIE, TFIIF, and TFIIH. The nucleating function of TFIID is thought to comprise several distinct events: (1) activator-dependent recognition of core promoter DNA elements; (2) the generation of a chromatin environment that is favorable to PlC assembly and transcription initiation; and (3) structural and functional modification of GTFs to facilitate PlC assembly and transcription initiation. Activities attributed to TAFII250 in vitro are consistent with roles for TAFII250 in each of these TFIID events, but evidence supporting these or other specific roles for TAFII250 in the context of chromatin is insufficient. We hypothesize that individual TAFII250 activities play gene-specific roles during RNA Pol II transcription and that signaling pathways and activators utilize individual TAFII250 activities to modulate the transcription of specific genes. Thus, in Aim 1 of the grant, we will use an unbiased genetic screen to identify TAFII250 domains and residues that are important for Drosophila viability. This screen will allow us to identify biologically relevant features of TAFII250. In Aim 2 of the grant, we will determine whether differential expression or differential chromatin association of TAFII250 C-terminal region (CTR) isoforms contributes to gene-specific transcription in Drosophila and whether TAFII250 features identified in Aim 1 contribute to this mechanism. In Aim 3 of the grant, we will determine whether differential biochemical activities of TAFII250 CTR isoforms contribute to gene-specific transcription in Drosophila and whether TAFII250 features identified in Aim 1 contribute to this mechanism. The determination of how individual TAFII250 activities contribute to the complex series of sequential interactions and conformational changes that promote transcription initiation in vivo will lead to a better understanding of mechanisms through which gene expression is regulated and how defects in this process lead to cancer and developmental abnormalities.

描述（由申请人提供）：TAFII250是与TFIID中与TBP复合的10个以上TATA结合蛋白（TBP）相关因子（TAFIIS）之一。 TFIID与围绕基因成核的转录起始位点的核心启动子的结合，其中包含RNA聚合酶II（POL II）和一般转录因子TFIIA，TFIIB，TFIIB，TFIIE，TFIIE，TFIIF和TFIIH。 TFIID的成核功能被认为包括几个不同的事件：（1）激活剂依赖性核心启动子DNA元素的识别； （2）有利于PLC组装和转录启动的染色质环境的产生； （3）GTF的结构和功能修饰，以促进PLC组装和转录启动。归因于TAFII250体外的活动与TFII250在每个TFIID事件中的角色一致，但是在染色质的背景下，支持TAFII250的这些或其他特定角色的证据不足。我们假设单个TAFII250活动在RNA POL II转录过程中起着基因特异性作用，并且信号通路和激活剂利用单个TAFII250活动来调节特定基因的转录。因此，在赠款的目标1中，我们将使用公正的遗传筛选来识别TAFII250域和对果蝇生存力很重要的残基。该屏幕将使我们能够识别TAFII250的生物学相关特征。在赠款的目标2中，我们将确定TAFII250 C末端区域（CTR）同工型的差异表达或差异染色质关联是否有助于果蝇中的基因特异性转录以及AIM 1中鉴定出的TAFII250特征是否有助于这种机制。在赠款的AIM 3中，我们将确定TAFII250 CTR同工型的差异化生物化学活性有助于果蝇中的基因特异性转录以及AIM 1中确定的TAFII250特征是否有助于这种机制。单个TAFII250活性如何促进一系列复杂的顺序相互作用和构象变化的确定，促进体内转录起始的构象变化将使人们更好地理解基因表达的机制，以及该过程中的缺陷如何导致癌症和发育异常。