title: Double-stranded DNA translocase activity of transcription factor TFIIH and the mechanism of RNA polymerase II open complex formation.
abstract: Formation of the RNA polymerase II (Pol II) open complex (OC) requires DNA unwinding mediated by the transcription factor TFIIH helicase-related subunit XPB/Ssl2. Because XPB/Ssl2 binds DNA downstream from the location of DNA unwinding, it cannot function using a conventional helicase mechanism. Here we show that yeast TFIIH contains an Ssl2-dependent double-stranded DNA translocase activity. Ssl2 tracks along one DNA strand in the 5' → 3' direction, implying it uses the nontemplate promoter strand to reel downstream DNA into the Pol II cleft, creating torsional strain and leading to DNA unwinding. Analysis of the Ssl2 and DNA-dependent ATPase activity of TFIIH suggests that Ssl2 has a processivity of approximately one DNA turn, consistent with the length of DNA unwound during transcription initiation. Our results can explain why maintaining the OC requires continuous ATP hydrolysis and the function of TFIIH in promoter escape. Our results also suggest that XPB/Ssl2 uses this translocase mechanism during DNA repair rather than physically wedging open damaged DNA.
authors.affiliation: Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98109; and.
authors.forename: James
authors.initials: J
authors.lastname: Fishburn
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Washington University in St. Louis, St. Louis, MO 63110.
authors.forename: Eric
authors.initials: E
authors.lastname: Tomko
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Washington University in St. Louis, St. Louis, MO 63110 egalburt@biochem.wustl.edu shahn@fhcrc.org.
authors.forename: Eric
authors.initials: E
authors.lastname: Galburt
authors.affiliation: Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98109; and egalburt@biochem.wustl.edu shahn@fhcrc.org.
authors.forename: Steven
authors.initials: S
authors.lastname: Hahn
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D001483:Base Sequence; D020134:Catalytic Domain; D004247:DNA / Q000737:chemistry; D004265:DNA Helicases / Q000378:metabolism*; D006868:Hydrolysis; D008969:Molecular Sequence Data; D011485:Protein Binding; D012319:RNA Polymerase II / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D013696:Temperature; D051758:Transcription Factor TFIIH / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: C489575:SSL2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D051758:Transcription Factor TFIIH; D000255:Adenosine Triphosphate; D004247:DNA; D012319:RNA Polymerase II; D000251:Adenosine Triphosphatases; D004265:DNA Helicases
keywords: DNA helicase; DNA unwinding; transcription initiation