title: Conformational flexibility and subunit arrangement of the modular yeast Spt-Ada-Gcn5 acetyltransferase complex.
abstract: The Spt-Ada-Gcn5 acetyltransferase (SAGA) complex is a highly conserved, 19-subunit histone acetyltransferase complex that activates transcription through acetylation and deubiquitination of nucleosomal histones in Saccharomyces cerevisiae. Because SAGA has been shown to display conformational variability, we applied gradient fixation to stabilize purified SAGA and systematically analyzed this flexibility using single-particle EM. Our two- and three-dimensional studies show that SAGA adopts three major conformations, and mutations of specific subunits affect the distribution among these. We also located the four functional modules of SAGA using electron microscopy-based labeling and transcriptional activator binding analyses and show that the acetyltransferase module is localized in the most mobile region of the complex. We further comprehensively mapped the subunit interconnectivity of SAGA using cross-linking mass spectrometry, revealing that the Spt and Taf subunits form the structural core of the complex. These results provide the necessary restraints for us to generate a model of the spatial arrangement of all SAGA subunits. According to this model, the chromatin-binding domains of SAGA are all clustered in one face of the complex that is highly flexible. Our results relate information of overall SAGA structure with detailed subunit level interactions, improving our understanding of its architecture and flexibility.
authors.affiliation: From the Department of Biochemistry and Molecular Biology, The University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada and.
authors.forename: Dheva
authors.initials: D
authors.lastname: Setiaputra
authors.affiliation: From the Department of Biochemistry and Molecular Biology, The University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada and.
authors.forename: James D
authors.initials: JD
authors.lastname: Ross
authors.affiliation: the National Institute of Biological Sciences, Beijing 102206, China.
authors.forename: Shan
authors.initials: S
authors.lastname: Lu
authors.affiliation: From the Department of Biochemistry and Molecular Biology, The University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada and.
authors.forename: Derrick T
authors.initials: DT
authors.lastname: Cheng
authors.affiliation: the National Institute of Biological Sciences, Beijing 102206, China.
authors.forename: Meng-Qiu
authors.initials: MQ
authors.lastname: Dong
authors.affiliation: From the Department of Biochemistry and Molecular Biology, The University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada and calvin.yip@ubc.ca.
authors.forename: Calvin K
authors.initials: CK
authors.lastname: Yip
mesh_terms: D000107:Acetylation; D002843:Chromatin / Q000737:chemistry* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D051548:Histone Acetyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006657:Histones / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation; D018583:Pliability; D011485:Protein Binding; D011487:Protein Conformation; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D054875:Ubiquitination
chemical_list: D002843:Chromatin; D006657:Histones; D021122:Protein Subunits; D029701:Saccharomyces cerevisiae Proteins; D051548:Histone Acetyltransferases
keywords: Acetyltransferase; Chromatin Modification; Deubiquitylation (Deubiquitination); Electron Microscopy (EM); Macromolecular Assembly; Mass Spectrometry (MS); Protein Cross-linking; Structural Biology; Yeast