title: The recruitment of the Saccharomyces cerevisiae Paf1 complex to active genes requires a domain of Rtf1 that directly interacts with the Spt4-Spt5 complex.
abstract: Transcription elongation factors associate with RNA polymerase II and aid its translocation through chromatin. One such factor is the conserved Paf1 complex (Paf1C), which regulates gene expression through several mechanisms, including the stimulation of cotranscriptional histone modifications. Previous studies revealed a prominent role for the Rtf1 subunit in tethering Paf1C to the RNA polymerase II elongation machinery. Here, we investigated the mechanism by which Rtf1 couples Paf1C to active chromatin. We show that a highly conserved domain of Rtf1 is necessary and sufficient for mediating a physical interaction between Rtf1 and the essential transcription elongation factor Spt5. Mutations that alter this Rtf1 domain or delete the Spt5 C-terminal repeat domain (CTR) disrupt the interaction between Rtf1 and Spt5 and release Paf1C from chromatin. When expressed in cells as the only source of Rtf1, the Spt5-interacting domain of Rtf1 can associate independently with active genes in a pattern similar to that of full-length Rtf1 and in a manner dependent on the Spt5 CTR. In vitro experiments indicate that the interaction between the Rtf1 Spt5-interacting domain and the Spt5 CTR is direct. Collectively, our results provide molecular insight into a key attachment point between Paf1C and the RNA polymerase II elongation machinery.
authors.affiliation: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
authors.forename: Manasi K
authors.initials: MK
authors.lastname: Mayekar
authors.forename: Richard G
authors.initials: RG
authors.lastname: Gardner
authors.forename: Karen M
authors.initials: KM
authors.lastname: Arndt
mesh_terms: D000595:Amino Acid Sequence; D002843:Chromatin / Q000378:metabolism; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D016366:Open Reading Frames; D054730:Protein Interaction Domains and Motifs; D060066:Protein Interaction Maps; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D016415:Sequence Alignment; D035181:TATA-Box Binding Protein / Q000737:chemistry / Q000378:metabolism*; D035602:Transcriptional Elongation Factors / Q000378:metabolism*
chemical_list: D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; D009687:Nuclear Proteins; C097698:PAF1 protein, S cerevisiae; C107273:RTF1 protein, S cerevisiae; C080991:SPT4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D035181:TATA-Box Binding Protein; D035602:Transcriptional Elongation Factors; C069940:SPT5 transcriptional elongation factor