Poster abstracts
Poster number 85 submitted by Le Luo
Mechanistic Insights into HIV-1 Tat Dependent Release of P-TEFb from the 7SK snRNP Complex
Le Luo (Department of Chemistry, Case Western Reserve University), Uri R. Mbonye (Department of Molecular Biology and Microbiology, School of Medicine, Case Western Reserve University), Jonathan Karn (Department of Molecular Biology and Microbiology, School of Medicine, Case Western Reserve University; Center for AIDS Research, Case Western Reserve University), Blanton S. Tolbert (Department of Chemistry, Case Western Reserve University; Center for AIDS Research, Case Western Reserve University)
Abstract:
The HIV-1 protein, Tat, stimulates transcriptional elongation by recruiting P-TEFb to the nascent RNAP II complex during transactivation of the proviral genome. P-TEFb exists in equilibrium between an inactive and active pool, where the 7SK snRNA along with the cellular protein HEXIM1 sequesters P-TEFb in its inactive state. The Tat protein is able to activate P-TEFb by releasing it from the inhibitory complex; however, the series of molecular events that promote the release mechanism is poorly understood. In this study, we demonstrate the specific binding of 7SK stem loops to HEXIM1 and Tat peptides using quantitative electrophoretic mobility shift assay. Both HEXIM1 and Tat can bind 7SK SL1 with nM affinities and HEXIM1 also associates with SL4 in the nanomolar range. Intriguingly, competition experiments between a Tat peptide and HEXIM1 for 7SK SL1 reveal Tat can effectively displace the HEXIM1-7SK complex. Thus, the results presented here offer insights into the mechanism used to promote the release of P-TEFb from the HEXIM1/7SK inhibitory complex.
Keywords: P-TEFb , HEXIM1-7SK, HIV-1 Tat