1. The Histone Modification Domain of Paf1 Complex Subunit Rtf1 Directly Stimulates H2B Ubiquitylation through an Interaction with Rad6.
- Author
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Van Oss SB, Shirra MK, Bataille AR, Wier AD, Yen K, Vinayachandran V, Byeon IL, Cucinotta CE, Héroux A, Jeon J, Kim J, VanDemark AP, Pugh BF, and Arndt KM
- Subjects
- Amino Acid Motifs, Binding Sites, Chromosomal Proteins, Non-Histone genetics, Chromosomal Proteins, Non-Histone metabolism, Cross-Linking Reagents chemistry, Crystallography, X-Ray, Formaldehyde chemistry, Histones chemistry, Histones genetics, Models, Molecular, Nuclear Proteins genetics, Nuclear Proteins metabolism, Protein Binding, Protein Conformation, alpha-Helical, Protein Interaction Domains and Motifs, RNA Polymerase II genetics, RNA Polymerase II metabolism, Recombinant Proteins chemistry, Recombinant Proteins genetics, Recombinant Proteins metabolism, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins chemistry, Saccharomyces cerevisiae Proteins genetics, TATA-Box Binding Protein chemistry, TATA-Box Binding Protein genetics, Transcription, Genetic, Transcriptional Elongation Factors genetics, Transcriptional Elongation Factors metabolism, Ubiquitin-Conjugating Enzymes chemistry, Ubiquitin-Conjugating Enzymes genetics, Ubiquitination, Gene Expression Regulation, Fungal, Histones metabolism, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins metabolism, TATA-Box Binding Protein metabolism, Ubiquitin-Conjugating Enzymes metabolism
- Abstract
The five-subunit yeast Paf1 complex (Paf1C) regulates all stages of transcription and is critical for the monoubiquitylation of histone H2B (H2Bub), a modification that broadly influences chromatin structure and eukaryotic transcription. Here, we show that the histone modification domain (HMD) of Paf1C subunit Rtf1 directly interacts with the ubiquitin conjugase Rad6 and stimulates H2Bub independently of transcription. We present the crystal structure of the Rtf1 HMD and use site-specific, in vivo crosslinking to identify a conserved Rad6 interaction surface. Utilizing ChIP-exo analysis, we define the localization patterns of the H2Bub machinery at high resolution and demonstrate the importance of Paf1C in targeting the Rtf1 HMD, and thereby H2Bub, to its appropriate genomic locations. Finally, we observe HMD-dependent stimulation of H2Bub in a transcription-free, reconstituted in vitro system. Taken together, our results argue for an active role for Paf1C in promoting H2Bub and ensuring its proper localization in vivo., (Copyright © 2016 Elsevier Inc. All rights reserved.)
- Published
- 2016
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