Mechanism of promoter melting by the xeroderma pigmentosum complementation group B helicase of transcription factor IIH revealed by protein-DNA photo-cross-linking

Mol Cell Biol. 2000 Nov;20(21):8168-77. doi: 10.1128/mcb.20.21.8168-8177.2000.

Abstract

The p89/xeroderma pigmentosum complementation group B (XPB) ATPase-helicase of transcription factor IIH (TFIIH) is essential for promoter melting prior to transcription initiation by RNA polymerase II (RNAPII). By studying the topological organization of the initiation complex using site-specific protein-DNA photo-cross-linking, we have shown that p89/XPB makes promoter contacts both upstream and downstream of the initiation site. The upstream contact, which is in the region where promoter melting occurs (positions -9 to +2), requires tight DNA wrapping around RNAPII. The addition of hydrolyzable ATP tethers the template strand at positions -5 and +1 to RNAPII subunits. A mutation in p89/XPB found in a xeroderma pigmentosum patient impairs the ability of TFIIH to associate correctly with the complex and thereby melt promoter DNA. A model for open complex formation is proposed.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Base Sequence
  • Blotting, Western
  • Cross-Linking Reagents
  • DNA / metabolism
  • Gene Deletion
  • HeLa Cells
  • Humans
  • Hydrolysis
  • Models, Biological
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Precipitin Tests
  • Promoter Regions, Genetic*
  • Protein Binding
  • Protein Structure, Tertiary
  • Proteins / metabolism
  • Recombinant Proteins / metabolism
  • Sequence Homology, Nucleic Acid
  • Transcription Factor TFIIH
  • Transcription Factors / genetics*
  • Transcription Factors, TFII*
  • Transcription, Genetic*
  • Xeroderma Pigmentosum / genetics*

Substances

  • Cross-Linking Reagents
  • Proteins
  • Recombinant Proteins
  • Transcription Factors
  • Transcription Factors, TFII
  • Transcription Factor TFIIH
  • Adenosine Triphosphate
  • DNA