The main goal of the proposed research is to understand the architecture, dynamics, and function of complex assemblies involved in transcriptional activation of human immunodeficiency virus type-1 (HIV-1) gene expression. HIV-1 encodes a transcriptional transactivator protein called Tat, which is expressed early in the viral life cycle and is absolutely required for viral replication and progression to disease. A regulatory element between +1 and +60 in the HIV-1 long terminal repeat which is capable of forming a stable stem-loop structure, designated TAR, is critical for Tat function. Tat interacts with cyclinT1 (CycT1), a regulatory partner of CDK9 in the positive transcription elongation factor b (P-TEFb) complex, and binds cooperatively with CycT1 to TAR RNA. Recruitment of P-TEFb to TAR promotes transcription elongation. P-TEFb is a key enzyme in the control of transcription elongation by RNA polymerase II and there are two pools of P-TEFb, active and inactive, present in the cell. P-TEFb is inactivated by sequestration into a large ribonucleoprotein (RNP) complex containing the small nuclear RNA, 7SK, and the Hexim1 protein. Therefore, there are two RNP complexes, TAR-Tat-P-TEFb and 7SK-Hexim1-P-TEFb, which are important in HIV-1 gene expression, however, it is not known how the equilibrium between these two RNPs is modulated. We have developed innovative chemical and physical approaches to probe RNA-protein and protein-protein interactions. The proposed work addresses the structure, dynamics, and function of TAR-Tat-P-TEFb and 7SK-Hexim1-P-TEFb complexes by revealing the molecular network of RNA-RNA and RNA-protein interactions that govern assembly and stability of the RNP complexes.
Results of these studies would contribute to understanding the mechanisms of assembly and stability of RNA-protein complexes under physiological conditions and will improve our understanding of HIV-1 gene regulationby Tat. Since P-TEFb is a key enzyme regulating cellular gene expression; understanding the mechanism of P-TEFb activation would provide fundamental insight into a number of regulatory processes involved in thedevelopment of diseases such as cardiac hypertrophy; cancer; inflammation; and autoimmunity. These resultswould also be valuable in developing new strategies for blocking the expression of retroviral or other disease-related genes.
Showing the most recent 10 out of 63 publications
