The long-term goal of the research described in this proposal is to elucidate fundamental aspects of HIV virion formation and RNA packaging. Although the outline of the process of assembly has been known for some time, and closely follows that of the simpler retroviruses, many unanswered questions remain. Elucidating the mechanism of HIV viral particle formation and RNA packaging in more detail is likely to help identify key events that could become targets for the next generation of anti-HIV therapeutics. The applicant will address the following specific questions:
Aim #1 : Do assembly and RNA packaging interactions begin on the polyribosome? The applicant will explore this question by expressing Gagpol together with Gag, and independently from it on a different mRNA, to determine whether sythesis of Gagpol by frameshifting contributes to its incorporation into particles. The applicant will also address the issue of cis versus trans packaging of viral RNA.
Aim #2 : Do RNA localization and processing mechanisms contribute to HIV RNA packaging? Although specific packaging signals are known to exist within HIV genomic RNA, and the NC protien seems to play role in RNA recognition, little is known about other factors that facilitate packaging. In this aim the applicant will: a) determine if poly A plays a role in RNA packaging; b) determine if targeting an RNA to the surface of the endoplasmic reticulum affects its ability to be packaged; c) determine if the pathway of nuclear export affects packaging efficiency; d) determine if newly identified RNA trafficking signals (RTS) present in the HIV genome and """"""""zipcodes"""""""" influence RNA packaging or assembly.
Aim #3 : What is the role of the 5' TAR element in RNA packaging and dimer linkage formation? When is the dimer linkage formed? The applicant has previously shown that the bottom part of the TAR stem is a necessary structural element for efficient RNA packaging. In this aim he will further explore this requirement, focusing attention on the possible role of TAR in dimer formation and dimer stability. He will also address the temporal order of dimer formation, i.e. can an RNA package without forming a dimer?
