[대학원 생명과학과 세미나 안내] 

연사 : 주유진 박사(Harvard Medical School)

연제 : Biochemical reconstitution system reveals a novel TAFs cofactor function for RNA polymerase II transcription

일시 : 2018년 4월 28일 (토) 오전 11시 

장소 : 하나과학관 A동 102호

초청교수 : 김준 교수

Abstract

RNA polymerase II (RNApII) binds many factors during the transcription process. These interactions must be clarified to understand how transcription is regulated. For this purpose, I sought to identify the RNApII complex components at each step of the transcription cycle. Through in vitro transcription reaction and quantitative mass spectrometry using immobilized templates, I was able to isolate the preinitiation complex on core promoter region and the promoter-driven RNApII elongation complex. These complexes contain basal factors, chromatin modulators, histone methyltransferases, and RNA processing factors that are already known to be involved in RNApII transcription. In addition, time course experiments revealed that RNApII C-terminal domain (CTD) phosphorylation and differential elongation complexes could be regulated by time, not a distance from transcription start site. Moreover, the binding factors dependent on CTD-phosphorylation could be identified by chemical inhibition of Kin28/Cdk7. Finally, experiments using a chromatin template confirmed that active transcription induced histone modifications including H3K36 and H3K79 methylation, as well as binding of FACT, Chd1, and ISWI chromatin remodelers to chromatin. These results help us in understanding the binding dynamics of various factors to the RNApII complex for proper gene expression. The post-transcriptional changes of TAFs (TBP-associated factor) interaction is a good example. TFIID has been known to bind promoter DNA to recruit RNA polymerase II and other basal factors for transcription. Although the TATA-binding protein (TBP) subunit of TFIID is necessary and sufficient for in vitro transcription, the TAF subunits recognize downstream promoter elements, act as coactivators, and interact with nucleosomes. In yeast nuclear extracts, transcription induces stable TAF binding to downstream promoter DNA, promoting subsequent activator-independent transcription reinitiation. In vivo, promoter responses to TAF mutations correlate with the level of downstream, rather than overall, Taf1 cross-linking. Based on these results, I propose a new model in which TAFs function as reinitiation factors, accounting for the differential responses of promoters to various transcription factor mutations.