The Retinoblastoma Tumor Suppressor Protein Targets Distinct General Transcription Factors To Regulate RNA Polymerase III Gene Expression

Abstract

The retinoblastoma protein (RB) represses RNA polymerase III transcription effectively both in vivo and in vitro. Here we demonstrate that the general transcription factors snRNA-activating protein complex (SNAP_c) and TATA binding protein (TBP) are important for RB repression of human U6 snRNA gene transcription by RNA polymerase III. RB is associated with SNAP_c as detected by both coimmunoprecipitation of endogenous RB with SNAP_c and cofractionation of RB and SNAP_c during chromatographic purification. RB also interacts with two SNAP_c subunits, SNAP43 and SNAP50. TBP or a combination of TBP and SNAP_crestores efficient U6 transcription from RB-treated extracts, indicating that TBP is also involved in RB regulation. In contrast, the TBP-containing complex TFIIIB restores adenovirus VAI but not human U6 transcription in RB-treated extracts, suggesting that TFIIIB is important for RB regulation of tRNA-like genes. These results suggest that different classes of RNA polymerase III-transcribed genes have distinct general transcription factor requirements for repression by RB.

ACKNOWLEDGMENTS

We are indebted to Nouria Hernandez for her generous support, helpful advice, and contribution of numerous reagents. We also gratefully thank Beicong Ma for constructing the GST-RB (379–928) expression plasmid and David Arnosti, Zachary Burton, and Craig Hinkley for critical reading of the manuscript.

This work was supported by grants from the American Cancer Society (RPG-00-263-01-GMC) and the Michigan State University Intramural Research Grant Program. Generous support was also provided by the Michigan State University College of Human Medicine and College of Natural Science.