Abstract
Retroviruses specifically package full-length, dimeric genomic RNA (gRNA) even in the presence of a vast excess of cellular RNA. The “psi” (Ψ) element within the 5′-untranslated region (5′UTR) of gRNA is critical for packaging through interaction with the nucleocapsid (NC) domain of Gag. However, in vitro Gag binding affinity for Ψ versus non-Ψ RNAs is not significantly different. Previous salt-titration binding assays revealed that human immunodeficiency virus type 1 (HIV-1) Gag bound to Ψ RNA with high specificity and relatively few charge interactions, whereas binding to non-Ψ RNA was less specific and involved more electrostatic interactions. The NC domain was critical for specific Ψ binding, but surprisingly, a Gag mutant lacking the matrix (MA) domain was less effective at discriminating Ψ from non-Ψ RNA. We now find that Rous sarcoma virus (RSV) Gag also effectively discriminates RSV Ψ from non-Ψ RNA in a MA-dependent manner. Interestingly, Gag chimeras, wherein the HIV-1 and RSV MA domains were swapped, maintained high binding specificity to cognate Ψ RNAs. Using Ψ RNA mutant constructs, determinants responsible for promoting high Gag binding specificity were identified in both systems. Taken together, these studies reveal the functional equivalence of HIV-1 and RSV MA domains in facilitating Ψ RNA selectivity by Gag, as well as Ψ elements that promote this selectivity.
Highlights
A hallmark of retroviruses is their ability to reverse transcribe single-stranded genomicRNA into double-stranded DNA for subsequent integration into the host cell genome.Following nuclear transcription and export into the cytoplasm, highly selective packaging of the full-length, unspliced genomic RNA (gRNA) into newly assembled virus particles is achieved, despite the presence of spliced viral RNA and a vast excess of cellular RNA [1,2]
These results suggest that Gag interacts with RNA using different binding modes; both the NC and MA domains bind to RNA in the case of TARpolyA, whereas binding to Ψ RNA preferentially involves the NC domain at the expense of MA interactions
We have shown that, as for Human immunodeficiency virus type 1 (HIV-1) Gag, specific recognition of Ψ RNA by Rous sarcoma virus (RSV) Gag involved a significantly greater nonelectrostatic binding component than interaction with non-Ψ RNA, and binding to Ψ was mediated primarily by the NC domain
Summary
A hallmark of retroviruses is their ability to reverse transcribe single-stranded genomicRNA (gRNA) into double-stranded DNA for subsequent integration into the host cell genome.Following nuclear transcription and export into the cytoplasm, highly selective packaging of the full-length, unspliced gRNA into newly assembled virus particles is achieved, despite the presence of spliced viral RNA and a vast excess of cellular RNA [1,2]. In addition to the NC domain, HIV-1 Gag consists of matrix (MA), capsid (CA), Viruses 2016, 8, 256; doi:10.3390/v8090256 www.mdpi.com/journal/viruses membrane (PM) [9], CA is involved in Gag multimerization associated with immature viral particle two spacer peptides, and p6 Inacid the context of the polyprotein, targets formation [10,11], NC functions as a nucleic chaperone in additionMA to its rolethe inassembling gRNA packaging virus to the plasma membrane (PM) [9], CA is involved in Gag multimerization associated with [12,13,14,15,16,17,18], and the p6 domain recruits factors required for viral fission from the cell [19,20]. Immature viral particle formation [10,11], NC functions as a nucleic acid chaperone in addition to its
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