A Nonconserved Surface of the TFIIB Zinc Ribbon Domain Plays a Direct Role in RNA Polymerase II Recruitment

Abstract

The general transcription factor TFIIB is a highly conserved and essential component of the eukaryotic RNA polymerase II (pol II) transcription initiation machinery. It consists of a single polypeptide with two conserved structural domains: an amino-terminal zinc ribbon structure (TFIIB_ZR) and a carboxy-terminal core (TFIIB_CORE). We have analyzed the role of the amino-terminal region of human TFIIB in transcription in vivo and in vitro. We identified a small nonconserved surface of the TFIIB_ZR that is required for pol II transcription in vivo and for different types of basal pol II transcription in vitro. Consistent with a general role in transcription, this TFIIB_ZR surface is directly involved in the recruitment of pol II to a TATA box-containing promoter. Curiously, although the amino-terminal human TFIIB_ZR domain can recruit both human pol II and yeast (Saccharomyces cerevisiae) pol II, the yeast TFIIB amino-terminal region recruits yeast pol II but not human pol II. Thus, a critical process in transcription from many different promoters—pol II recruitment—has changed in sequence specificity during eukaryotic evolution.

We thank L. Freedman, P. Hearing, N. Hernandez, and members of the Herr laboratory for helpful discussions and suggestions during the course of this work; H. Ge and N. Kaludov (ProteinOne, Inc.) for technical advice; D. Reinberg and S. Mandal for highly purified human pol II and TFIIF antibody 6H10; R. Kornberg and D. Bushnell for purified S. cerevisiae pol II; D. Aufiero for recombinant baculovirus-expressed SNAP_C; N. Hernandez and T. Kuhlman for the α-IIB/4 antiserum; S. Hahn for the pLB2SUA7 S. cerevisiae TFIIB construct; M. Wang for help with structural modeling; N. Hernandez, E. Julien, and X. Zhao for helpful comments on the manuscript; and J. Duffy for artwork.

These studies were supported by Public Health Service grant CA13106.