Poster abstracts

Poster number 23 submitted by Andrew Goodrich

Regulation of reverse transcription by HIV-1 Gag

Andrew C. Goodrich (Departments of Chemistry and Biochemistry, The Ohio State University), Christopher P. Jones (Departments of Chemistry and Biochemistry, The Ohio State University), Karin Musier-Forsyth (Departments of Chemistry and Biochemistry, The Ohio State University)

Abstract:
Human immunodeficiency virus type-1 (HIV-1) utilizes human cellular tRNA^Lys in a critical step of its life cycle—the priming of reverse transcription of its single-stranded RNA into double-stranded DNA. For priming to occur, tRNA^Lys must be annealed to the complementary 18-nucleotide site known as the Primer Binding Site (PBS) on the viral genome. Biological evidence suggests this process is performed by the viral Gag protein, the structural protein responsible for viral assembly. During the viral life cycle, Gag is proteolyzed into the matrix (MA), capsid (CA), nucleocapsid (NC), and p6 proteins. A potent nucleic acid chaperone, NC plays a major role in restructuring RNA and DNA throughout the reverse transcription process. NC has been shown to anneal the tRNA^Lys primer by promoting rearrangement of the PBS:tRNA^Lys duplex to the thermodynamically most stable conformation. NC consists of two “zinc finger” motifs flanked by unstructured basic residues. Although both NC and Gag facilitate tRNA primer annealing to the PBS in vitro, we find reverse transcriptase extension is inhibited in the presence of Gag. Interestingly, in vivo studies have shown that Gag variants with zinc finger deletions or point mutations undergo premature reverse transcription, abolishing infectivity. In this work, similar Gag mutants were tested for their ability to bind and aggregate nucleic acids and to facilitate tRNA annealing and reverse transcription in vitro. We show that the nucleic acid chaperone activity of these Gag variants is robust and propose an explanation for Gag’s role in inhibiting premature reverse transcription.

Keywords: HIV-1, reverse transcription, RNA chaperone