Transcription elongation through DNA arrest sites. A multistep process involving both RNA polymerase II subunit RPB9 and TFIIS.
The role of yeast RNA polymerase II (pol II) subunit RPB9 in transcript elongation was investigated by examining the biochemical properties of pol II lacking RPB9 (pol IIDelta9). The maximal rate of chain elongation was nearly identical for pol II and pol IIDelta9. By contrast, pol IIDelta9 elongated more efficiently through DNA sequences that signal the elongation complex to pause or arrest. The addition of purified recombinant RPB9 to pol IIDelta9 restored the elongation properties of the mutant polymerase to those of the wild-type enzyme. Arrested pol IIDelta9 complexes were refractory to levels of TFIIS that promoted maximal read-through with pol II. However, both pol II and pol IIDelta9 complexes stimulated with TFIIS undergo transcript cleavage, confirming that transcript cleavage and read-through activities can be uncoupled. Our observations suggest that both TFIIS and RPB9 are required to stimulate the release of RNA polymerase II from the arrested state.
Awrey
D E
DE
Cancer Research Group, Institute for Molecular Biology and Biotechnology, McMaster University, Hamilton, Ontario L8N 3Z5, Canada.
Weilbaecher
R G
RG
Hemming
S A
SA
Orlicky
S M
SM
Kane
C M
CM
Edwards
A M
AM
D003001:Cloning, Molecular; D017353:Gene Deletion; D005982:Glutathione Transferase; D007700:Kinetics; D012319:RNA Polymerase II / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D035165:Transcription Factors, General; D014158:Transcription, Genetic; D035602:Transcriptional Elongation Factors
D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D035165:Transcription Factors, General; D035602:Transcriptional Elongation Factors; C018648:transcription factor S-II; D005982:Glutathione Transferase; D012319:RNA Polymerase II