YEAST GENES INVOLVED IN GENERAL TRANSCRIPTIONAL CONTROL

参与一般转录控制的酵母基因

• 批准号: 6180117
• 负责人: CLYDE L DENIS
• 金额: $ 36.6万
• 依托单位: UNIVERSITY OF NEW HAMPSHIRE
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-09-01 至 2003-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6180117
• 关键词: cell cycle; fungal genetics; gene expression; genetic models; genetic regulation; genetic regulatory element; genetic transcription; mass spectrometry; protein protein interaction; protein purification; protein structure function; transcription factor

The goal of this research is to elucidate the mechanisms of eucaryotic gene regulation. The system chosen to study is the multi-protein CCR4-NOT complex, which affects gene expression both positively and negatively. It consists of two complexes, 1.9 and 1.0 mD in size, that are distinct from other transcriptionally important groups of proteins, such as the SNF/SWI and SAGA complexes, TFIID, and the RNA polymerase II holoenzyme. The smaller of the CCR4-NOT complexes contains CCR4, CAF1, the five NOT proteins and several unidentified proteins. The repressive effects of the CCR4-NOT proteins appear to occur through destabilizing TBP association with non-canonical TATAA sequences, probably through their direct interaction with TFIIA, TAFs or other TBP-associated factors. The SRB9-11 holoenzyme subcomplex also displays physical and functional interactions with the CCR4-NOT complex. The central focus of this grant proposal is three-fold: 1) What are the components and physiological roles of the two CCR4-NOT complexes? 2) How do the CCR4-NOT proteins affect the function of TBP and its associated factors? and 3) How is the CCR4-NOT complex required for activated transcription? The 1.9 mD CCR4-NOT complex will be isolated, its components cloned following mass spectrometric analysis, and their functions characterized. The mechanism by which the CCR4-NOT complex affects the action of TBP and its associated factors will be elaborated by several biochemical studies. Genetic screens will be undertaken to identify new factors or novel interactions between the CCR4-NOT complex and transcriptionally important proteins. CCR4 activator function will be explored through characterizing its unique C- and N-terminal interactions with SRB9-11, SPT5, TAF60, and other proteins. The key features of the ADH2 promoter which are responsive to the CCR4-NOT complex will be identified to determine how the CCR4-NOT complex controls the ADH2 gene in a positive manner.

这项研究的目的是阐明桉树基因调节的机制。 选择研究的系统是多蛋白CCR4-不复杂的复合物，它既积极和负面影响基因表达。 它由两种络合物组成：1.9和1.0 MD的大小，它们与其他具有转录重要的蛋白质组不同，例如SNF/SWI和SAGA复合物，TFIID和RNA聚合酶II Holoenzyme。 较小的CCR4-NOT复合物包含CCR4，CAF1，五种非蛋白质和几种未鉴定的蛋白质。 CCR4-NOT蛋白的抑制作用似乎是通过使TBP与非典型TATAA序列的稳定稳定发生的，这可能是通过与TFIIA，TAFS或其他与TBP相关因素的直接相互作用而发生的。 SRB9-11 Holoenzyme子复合物还显示了与CCR4-NOT复合物的物理和功能相互作用。 该赠款提案的主要重点是三重：1）两个CCR4不复合物的组成部分和生理作用是什么？ 2）CCR4不蛋白如何影响TBP及其相关因素的功能？ 3）如何激活转录需要CCR4不合适的复合物？ 1.9 MD CCR4不复合物将被分离出来，其组件在质谱分析后克隆，其功能表征。 CCR4-不复合物影响TBP的作用及其相关因素的机制将通过多项生化研究阐述。 将进行遗传筛选，以确定CCR4不复合物与转录重要蛋白质之间的新因素或新型相互作用。 CCR4激活函数将通过表征其独特的C和N末端相互作用与SRB9-11，SPT5，TAF60和其他蛋白质进行探索。 将确定对CCR4（不反应CCR4）的ADH2启动子的关键特征，以确定CCR4-NOT复合物如何以阳性方式控制ADH2基因。