Analysis of General Transcription Factors in Yeast

这个应用程序的长期目标是了解更多关于基本和保守的 真核生物中转录起始的方面。这些研究将集中在一种蛋白质复合物上， 一种酵母，酿酒酵母，通过RNA聚合酶I 1控制转录起始。这 复合物，命名为佐贺（Spt-Ada-Gcn 5乙酰转移酶），是一类因子的成员，称为 辅激活因子，在转录起始的控制中起关键作用。辅活化剂是大的，多- 蛋白质复合物通常具有多种活性，在某些情况下可以激活和抑制 转录。佐贺是保守的，因为已经鉴定了人SAGA样复合物。若干问题 关于共激活剂的了解很少，包括它们的激活机制，功能 不同共激活因子之间的冗余，以及一些共激活因子如何既激活又抑制 转录。 本实验旨在研究佐贺在酵母中的功能。具体 目标1包括三组实验。前两组解决了激活机制 佐贺组件，Spt 3，使用生化和遗传方法。第三组解决了可能的 一些佐贺亚基乙酰化的作用。具体目标2解决了佐贺的体内协调 与其他辅活化剂复合物的活性。第一部分将基因分析与微阵列相结合 研究，以解决佐贺和第二辅活化剂复合物，Swi/Snf之间的功能冗余。 第二部分旨在鉴定与佐贺一起激活转录的其他因子。 研究充分的GAL 1启动子。具体目标3将焦点转移到解决转录抑制。Mot 3是 一种与佐贺功能相关的DNA结合阻遏物。Mot 3在抑制细胞凋亡中起着关键作用。 麦角甾醇是S.酿酒酵母膜。强镇压 也需要低氧水平。提出了两组实验。一是 抑制机制将得到解决，测试的初步发现，佐贺也需要 抑制这些基因，确定其他必要的因素，并阐明其各自的作用。 第二，氧介导的调节作用将使用分子和遗传分析来解决。 这些研究应该揭示在酵母转录控制的重要方面。考虑到强劲的 保守的转录机制，结果将适用于理解转录， 人类

The long-term objectives of this application are to learn more about fundamental and conserved aspects of transcription initiation in eukaryotes. The studies will focus on a protein complex discovered in the yeast, Saccharomyces cerevisiae, that controls transcription initiation by RNA polymerase I1. This complex, named SAGA (Spt-Ada-Gcn5 Acetyltransferase), is a member of a class of factors, called coactivators, that play critical roles in the control of transcription initiation. Coactivators are large, multi- protein complexes that often possess multiple activities and in some cases can both activate and repress transcription. SAGA is conserved, as human SAGA-like complexes have been identified. Several issues about coactivators are poorly understood, including their mechanisms of activation, the functional redundancy between different coactivators, and how some coactivators can both activate and repress transcription. The proposed experiments are to study several issues concerning SAGA's functions in yeast. Specific Aim 1 contains three sets of experments. The first two sets address the mechanism of activation by the SAGA component, Spt3, using biochemical and genetic approaches. The third set addresses the possible role of acetylation of some SAGA subunits. Specific Aim 2 addresses the in vivo coordination of SAGA's activities with other coactivator complexes. The first section combines genetic analysis with microarray studies to address the functional redundancy between SAGA and a second coactivator complex, Swi/Snf. The second section is designed to identify other factors that function with SAGA to activate transcription of the well-studied GAL 1 promoter. Specific Aim 3 shifts focus to address repression of transcription. Mot3 is a DNA-binding repressor that is functionally related to SAGA. Mot3 plays a critical role in the repression of genes required for synthesis of ergosterol, a key component of S. cerevisiae membranes. Strong repression of these genes also requires low oxygen levels. Two sets of experiments are proposed. First, the mechanism of repression will be addressed, testing the preliminary finding that SAGA is also required for repression of these genes, identifying other factors necessary, and elucidating their respective roles. Second, the role of oxygen-mediated regulation will be addressed using molecular and genetic analysis. These studies should reveal important aspects of transcriptional control in yeast. Given the strong conservation of transcriptional mechanisms, the results will be applicable to understanding transcription in humans.