Analysis of the mechanism of eukaryotic transcription initiation and its regulation based on the TATA box-binding factor TFIID

TATA盒结合因子TFIID在转录激活机制中起着至关重要的作用。我们通过多种方法阐明了TFIID在真核转录启动中的功能和结构的意义及其调控。1)编码人TFIID亚单位的cDNA的分离和鉴定2)TFIID内的组蛋白八聚体样结构模型我们以其他与组蛋白H4相似的TFIID亚基为探针，分离出结构类似于组蛋白H_2B的TFIID亚基，表明TFIID亚基之间的相互作用是由各自的组蛋白折叠基序引起的。3)从几个物种中分离到编码TFIID亚基的基因。为了阐明TFIID亚基的结构与功能之间的关系，我们从几个物种中分离了TFIID亚基的编码基因。4)TFIID亚基的遗传学分析为了阐明TFIID亚基在体内的功能，我们从几个物种中分离了TFIID亚基。发展了一种分离温度敏感突变体的新方法。分离的ts突变体TFIID亚基存在胞质分裂缺陷。5)分离编码与TFIID亚基相互作用的各种新因子的cDNA。这些孤立的因素大多与染色质有关。其中一些被证明具有组蛋白乙酰转移酶活性和核小体重塑活性。

TATA box-binding factor TFIID plays a crucial role in machanisms of transcription activation.We elucidated the significance of function and structure of TFIID in eukaryotic transcription initiation and its regulation by several methods.1)Isolation and characterization of cDNA encoding human TFIID subunitsTwo novel types of cDNAs, which encode human TFIID subunits and have similalities to histones, were isolated. The analysis of interactions between these subunits and other TFIID subunits has been made.2)Model of histone octamer-like structure within TFIIDWe isolated TFIID subunit whose structure is similar to that of histone H2B using other TFIID subunit with histone H4 similarity as a probe and showed that the interaction of two subunits is caused by their own histone folding motifs. We also isolated other TFIID subunit similar to histone H2A and showed a model of histone octomer-like structure.3)Isolation of cDNA encoding TFIID subunits from several speciesWe isolated cDNA encoding TFIID subunits from several species in order to elucidate the relationship between structure and function.4)Genetical analysis of S.pombe TFIID subunitsTo elucidate the in vivo function of TFIID subunits, we isolated S.pombe TFIID subunits. New method for isolation of temparature sensitive mutants are developed. Isolated ts mutant TFIID subunit shows defect in cytokinesis.5)Isolation of cDNAs encoding various new factors interacting with TFIID subunitscDNAs encoding various new factors interacting with TFIID subunits were isolated. Most of these isolated factors are chromatin-related. Some of them are shown to have histone acetyl transferase activities and nucleosome-remodeling activities.