Studies on the regulatory mechanisms of TFIID-mediated eukaryotic transcription

酿酒酵母（Saccharomyces cerevisiae） HM01是一种高迁移率的B组蛋白（HMGB），与rRNA位点和许多核糖体蛋白基因（rpg）的启动子相关。本研究利用Sos招募系统证明了HM01与TBP和TAF1的n端结构域（TAND）相互作用，它们是TFIID的组成部分。生化研究表明，HM01与TFIID发生协同作用，并与TBP直接相互作用，而不与TAND相互作用。缺失HM01 （Deltahmol）会导致严重的冷敏感生长缺陷，并降低一些tand依赖基因的转录。Deltahmol也以特定于某些RPG启动子的方式影响TFIID占用。有趣的是，过表达HM01会延迟TAND缺失突变体（taf1DeltaTAND）的集落形成，而野生型菌株则不会，这表明HM01与TAND之间存在功能联系。此外，Deltahmol在某些spt15 （TBP）和total （TFIIA）突变体中表现出合成生长缺陷，而在某些su…More a7 （TFIIB）突变体中却能恢复生长缺陷。重要的是，Deltahmol导致RPS5、RPS16A、RPL23B、RPL27B和RPL32的转录起始位点上游移动，但不影响RPS31、RPL10、TEF2和ADH1的转录起始位点，这表明HM01可能通过与TFIID的相互作用参与了II类基因子集的起始位点选择。本研究还利用全基因组染色质免疫沉淀技术研究了HMO1、FHL1和RAP1在RPG转录中的作用。结果表明，基于启动子上HM01的丰度以及FHL1和/或RAP1与启动子结合对HM01的依赖性，RPG启动子（共138个）可分为几个不同的组。FHL1以依赖hmo1的方式结合大多数hmo1富集和转录依赖于hmo1的RPG启动子，而无论它们是否以依赖hmo1的方式转录，FHL1都以不依赖hmo1的方式结合hmo1受限的RPG启动子。重要的是，这些功能特性是由启动子序列决定的。少

Saccharomyces cerevisiae HM01, a high mobility group B (HMGB) protein, associates with the rRNA locus and with the promoters of many ribosomal protein genes (RPGs) .Here, the Sos recruitment system was used to show that HM01 interacts with TBP and the N-terminal domain (TAND) of TAF1, which are integral components of TFIID. Biochemical studies revealed that HM01 copurifies with TFIID and directly interacts with TBP but not with TAND. Deletion of HM01 (Deltahmol) causes a severe cold-sensitive growth defect and decreases transcription of some TAND-dependent genes. Deltahmol also affects TFIID occupancy at some RPG promoters in a promoter-specific manner. Interestingly, over-expression of HM01 delays colony formation of tafl mutants lacking TAND (taf1DeltaTAND), but not of the wild-type strain, indicating a functional link between HM01 and TAND. Furthermore, Deltahmol exhibits synthetic growth defects in some spt15 (TBP) and toal (TFIIA) mutants while it rescues growth defects of some su … More a7 (TFIIB) mutants. Importantly, Deltahmol causes an upstream shift in transcriptional start sites of RPS5, RPS16A, RPL23B, RPL27B and RPL32, but not of RPS31, RPL10, TEF2 and ADH1, indicating that HM01 may participate in start site selection of a subset of class II genes presumably via its interaction with TFIID.This study also uses genome-wide chromatin immunoprecipitation to study the roles of HMO1, FHL1, and RAP1 in RPG transcription. The results show that RPG promoters(138 in total) can be classified into several distinct groups based on HM01 abundance at the promoter and the HM01 dependence of FHL1 and/or RAP1 binding to the promoter. FHL1 binds to most of the HMO1-enriched and transcriptionally HMO1-dependent RPG promoters in an HMO1-dependent manner, whereas it binds to HMO1-limited RPG promoters in an HMO1-independent manner, irrespective of whether they are transcribed in an HMO1-dependent manner. Importantly, these functional properties are determined by the promoter sequence. Less