title: Insights into how Spt5 functions in transcription elongation and repressing transcription coupled DNA repair.
abstract: Spt5, a transcription elongation factor, and Rpb4, a subunit of RNA polymerase II (RNAP II) that forms a subcomplex with Rpb7, play important roles in transcription elongation and repression of transcription coupled DNA repair (TCR) in eukaryotic cells. How Spt5 physically interacts with RNAP II, and if and/or how Spt5 and Rpb4/7 coordinate to achieve the distinctive functions have been enigmatic. By site-specific incorporation of the unnatural amino acid p-benzoyl-L-phenylalanine, a photoreactive cross-linker, we mapped interactions between Spt5 and RNAP II in Saccharomyces cerevisiae. Through its KOW4-5 domains, Spt5 extensively interacts with Rpb4/7. Spt5 also interacts with Rpb1 and Rpb2, two largest subunits of RNAP II, at the clamp, protrusion and wall domains. These interactions may lock the clamp to the closed conformation and enclose the DNA being transcribed in the central cleft of RNAP II. Deletion of Spt5 KOW4-5 domains decreases transcription elongation and derepresses TCR. Our findings suggest that Spt5 is a key coordinator for holding the RNAP II complex in a closed conformation that is highly competent for transcription elongation but repressive to TCR.
authors.affiliation: Department of Comparative Biomedical Sciences, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA, USA.
authors.forename: Wentao
authors.initials: W
authors.lastname: Li
authors.affiliation: Department of Biological Sciences, Louisiana State University, Baton Rouge, LA, USA.
authors.forename: Cristina
authors.initials: C
authors.lastname: Giles
authors.affiliation: Department of Comparative Biomedical Sciences, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA, USA shli@vetmed.lsu.edu.
authors.forename: Shisheng
authors.initials: S
authors.lastname: Li
mesh_terms: D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry / Q000378:metabolism*; D004260:DNA Repair; D009687:Nuclear Proteins / Q000737:chemistry; D054730:Protein Interaction Domains and Motifs; D012319:RNA Polymerase II / Q000737:chemistry* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D061805:Transcription Elongation, Genetic; D019473:Transcription Factors, TFII / Q000378:metabolism; D024363:Transcription Initiation Site; D014158:Transcription, Genetic; D035602:Transcriptional Elongation Factors / Q000737:chemistry / Q000378:metabolism*
chemical_list: D002868:Chromosomal Proteins, Non-Histone; D009687:Nuclear Proteins; C080991:SPT4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D019473:Transcription Factors, TFII; D035602:Transcriptional Elongation Factors; C066644:transcription factor TFIIE; C069940:SPT5 transcriptional elongation factor; D012319:RNA Polymerase II; C423309:RPB4 protein, S cerevisiae; C470553:RPB7 protein, S cerevisiae