Simultaneous recruitment of coactivators by Gcn4p stimulates multiple steps of transcription in vivo.
Transcriptional activation by Gcn4p is dependent on the coactivators SWI/SNF, SAGA, and Srb Mediator, which are recruited by Gcn4p and stimulate assembly of the pre-initiation complex (PIC) at the ARG1 promoter in vivo. We show that recruitment of all three coactivators is nearly simultaneous with binding of Gcn4p at ARG1 and is followed quickly by PIC formation and elongation by RNA polymerase II (Pol II) through the open reading frame. Despite the simultaneous recruitment of coactivators, rapid recruitment of SWI/SNF depends on the histone acetyltransferase (HAT) subunit of SAGA (Gcn5p), a non-HAT function of SAGA, and on Mediator. SAGA recruitment in turn is strongly stimulated by Mediator and the RSC complex. Recruitment of Mediator, by contrast, occurs independently of the other coactivators at ARG1. We confirm the roles of Mediator and SAGA in TATA binding protein (TBP) recruitment and demonstrate that all four coactivators under study enhance Pol II recruitment or promoter clearance following TBP binding. We also present evidence that SWI/SNF and SAGA stimulate transcription elongation downstream from the promoter. These functions can be limited to discrete time intervals, providing evidence for multiple stages in the induction process. Our findings reveal a program of coactivator recruitment and PIC assembly that distinguishes Gcn4p from other yeast activators studied thus far.
Govind
Chhabi K
CK
Laboratory of Gene Regulation and Development, National Institute of Child Health and Human Development, Bethesda, Maryland 20892, USA.
Yoon
Sungpil
S
Qiu
Hongfang
H
Govind
Sudha
S
Hinnebusch
Alan G
AG
D000123:Acetyltransferases / Q000378:metabolism; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D051548:Histone Acetyltransferases; D007700:Kinetics; D008954:Models, Biological; D009154:Mutation; D016366:Open Reading Frames; D011401:Promoter Regions, Genetic; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D021122:Protein Subunits / Q000378:metabolism; D012319:RNA Polymerase II / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D035381:TATA-Binding Protein Associated Factors / Q000235:genetics / Q000378:metabolism; D035181:TATA-Box Binding Protein / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D004268:DNA-Binding Proteins; D021122:Protein Subunits; C103337:RSC complex, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D035381:TATA-Binding Protein Associated Factors; D035181:TATA-Box Binding Protein; D015534:Trans-Activators; D014157:Transcription Factors; D000123:Acetyltransferases; D051548:Histone Acetyltransferases; D011494:Protein Kinases; D012319:RNA Polymerase II