Tat-SF1 Protein Associates with RAP30 and Human SPT5 Proteins

Abstract

The potent transactivator Tat recognizes the transactivation response RNA element (TAR) of human immunodeficiency virus type 1 and stimulates the processivity of elongation of RNA polymerase (Pol) II complexes. The cellular proteins Tat-SF1 and human SPT5 (hSPT5) are required for Tat activation as shown by immunodepletion with specific sera and complementation with recombinant proteins. In nuclear extracts, small fractions of both hSPT5 and Pol II are associated with Tat-SF1 protein. Surprisingly, the RAP30 protein of the heterodimeric transcription TFIIF factor is associated with Tat-SF1, while the RAP74 subunit of TFIIF is not coimmunoprecipitated with Tat-SF1. Overexpression of Tat-SF1 and hSPT5 specifically stimulates the transcriptional activity of Tat in vivo. These results suggest that Tat-SF1 and hSPT5 are indispensable cellular factors supporting Tat-specific transcription activation and that they may interact with RAP30 in controlling elongation.

ACKNOWLEDGMENTS

We thank E. Lees, D. Reinberg, and R. Gaynor for valuable materials and T. P. Cujec and B. M. Peterlin for providing the Tat-HA expression vector. We thank A. Mitsui, S. Gilbert, D. Dykxhoorn, D. Tantin, and H. Tang for valuable advice and helpful comments. We also thank M. Siafaca for secretarial support.

This work was supported by U.S. Public Health Service grants RO1-AI32486 and PO1-CA42063 from the National Institutes of Health, by NCI Cancer Center Support (core) grant P30-CA 14051 to P.A.S., by a Grant-in-Aid for Scientific Research on Priority Areas from the Ministry of Education, Science, Sports and Culture of Japan, and by a research grant from CREST of JST Corporation to H.H. J.B.K. was supported by an Anna-Fuller postdoctoral fellowship. Y.Y. was a JSPS research fellow.