The HIV life cycle requires the export of intron-containing viral mRNAs from the nucleus to the cytoplasm. This process is ordinarily restricted by the cell, but HIV overcomes this check through the interaction of a viral protein, Rev, and an RNA secondary structure, the Rev-Response Element (RRE), found on intron-containing viral transcripts. Variation in the functional activity of the Rev-RRE axis has been observed in viruses taken from patients infected with HIV, but the clinical significance of that variation and its causal mechanism is not clearly understood. Previous studies of a related lentivirus clearly show that Rev-RRE activity variation plays a role in pathogenesis, strongly suggesting that this system is similarly important in HIV infection. It is hypothesized that the Rev-RRE regulatory axis functions as a rheostat which permits viral adaptation to different immune environments. In this K08 Mentored Clinical Scientist Research Career Development Award application, Dr. Patrick Jackson, a fellow in Infectious Diseases at the University of Virginia, proposes to perform a series of studies to illuminate the role of the Rev-RRE system in HIV transmission and in viral latency. First, using a novel high-throughput lentiviral vector assay of Rev-RRE functional activity, the role of this axis in HIV transmission will be examined by comparing Rev-RRE activity in transmitted and circulating viral quasispecies. Second, the Rev-RRE activity of latent proviruses and circulating viruses will be compared. High and low activity Rev-RRE pairs will be tested to see if they cause differences in viral replication kinetics and the production of viral proteins. This phenomenon could underlie the failure of ?kick and kill? strategies to cure HIV. Finally, domain swapping experiments will be performed to identify the sequence determinants of both Rev and RRE functional activity. Naturally occurring and chimeric Rev and RRE sequences will be utilized to identify key residues and structures. This research has direct implications for the development of improved pharmacologic prophylaxis for HIV transmission and for improvements in ?kick and kill? HIV cures. This research will be conducted at the University of Virginia under the mentorship of Drs. David Rekosh and Marie- Louise Hammarskjold who have a strong track record of training success. Dr. Jackson proposes a career development plan consisting of experiential learning; formal course work in molecular biology, bioinformatics, and grant writing; and structured research and professional mentorship. He proposes a specific timeline for presentation of results at international conferences and in peer reviewed publications, as well as a timeline for future grant applications. This program is designed to situate him as an independent researcher with expertise in HIV virology and a skill set which encompasses a broad set of scientific techniques.
. The HIV Rev-RRE regulatory system is a key step in the virus life cycle, investigation of which will yield insights into both host and pathogen post-transcriptional gene regulation. Improved understanding of this system will have significance for understanding HIV transmission and viral latency, and will contribute to the development of improved pharmacologic prophylaxis and ?kick and kill? strategies to cure HIV.