Mechanisms of P-TEFb Regulation and Recruitment by Tat

Seemay Chou, University of California Berkeley
Advisor: Tom Alber
Training in Basic Biomedical Sciences
Dissertation Award

Through direct interactions with the viral hairpin RNA, TAR, as well as host transcriptional machinery and transcriptional regulators, HIV Tat enhances elongation of HIV transcripts. Tat binds the positive transcription elongation factor b (P-TEFb) component, cyclin T1, as well as TAR in order to bring the P-TEFb complex to the HIV-1 promoter where it phosphorylates and stimulates elongating RNA polymerase II while inhibiting negative elongation factors. Tat may also allosterically activate P-TEFb by stimulating Cdk9 kinase activity.

Tat also increases the total population of active P-TEFb population. Cellular P-TEFb exists in both an inactive and active state, and Tat can shift this carefully regulated equilibrium to upregulate P-TEFb activity. Inactive P-TEFb exists in a large complex with 7SK snRNA as well as several proteins, including HEXIM1, PIP7S, and BCDIN3. In contrast, active P-TEFb is bound by the bromodomain-containing protein Brd4, which stimulates host transcription. Tat not only releases P-TEFb from the inhibitory 7SK snRNP complex, but also competes with Brd4 for P-TEFb recruitment to the HIV promoter.

Tat-dependent transactivation is critical for successful HIV transcription. Disruption of Tat or P-TEFb results in the production of abortive HIV transcripts. However, the molecular mechanisms by which Tat modulates P-TEFb complex activity remain unclear. Structurally defining the interactions between Tat and host proteins may provide insight into Tat mechanism. Moreover, preliminary studies suggest that additional unidentified components of these complexes may be regulated by Tat or required for Tat transactivation. Therefore, the biochemical and structural characterization of P-TEFb complexes is necessary to better understanding Tat function and uncover novel therapeutic targets.