Abstract
How retroviruses regulate the amount of RNA genome packaged into each virion has remained a long-standing question. Our previous study showed that most HIV-1 particles contain two copies of viral RNA, indicating that the number of genomes packaged is tightly regulated. In this report, we examine the mechanism that controls the number of RNA genomes encapsidated into HIV-1 particles. We hypothesize that HIV-1 regulates genome packaging by either the mass or copy number of the viral RNA. These two distinct mechanisms predict different outcomes when the genome size deviates significantly from that of wild type. Regulation by RNA mass would result in multiple copies of a small genome or one copy of a large genome being packaged, whereas regulation by copy number would result in two copies of a genome being packaged independent of size. To distinguish between these two hypotheses, we examined the packaging of viral RNA that was larger (≈17 kb) or smaller (≈3 kb) than that of wild-type HIV-1 (≈9 kb) and found that most particles packaged two copies of the viral genome regardless of whether they were 17 kb or 3 kb. Therefore, HIV-1 regulates RNA genome encapsidation not by the mass of RNA but by packaging two copies of RNA. To further explore the mechanism that governs this regulation, we examined the packaging of viral RNAs containing two packaging signals that can form intermolecular dimers or intramolecular dimers (self-dimers) and found that one self-dimer is packaged. Therefore, HIV-1 recognizes one dimeric RNA instead of two copies of RNA. Our findings reveal that dimeric RNA recognition is the key mechanism that regulates HIV-1 genome encapsidation and provide insights into a critical step in the generation of infectious viruses.
Highlights
Retroviruses are RNA viruses that replicate through a DNA phase, in which viral DNA is integrated into the host genome to form a provirus [1]
Many aspects of RNA packaging are well-studied, how retroviruses regulate the number of genomes in the particle is currently unknown
This assumption was validated recently when we demonstrated that most HIV-1 particles contain two copies of viral RNA, which revealed that the number of genomes packaged is tightly controlled
Summary
Retroviruses are RNA viruses that replicate through a DNA phase, in which viral DNA is integrated into the host genome to form a provirus [1]. Packaging of the retroviral genome is mediated by interactions between the viral structural protein Gag and the cis-acting element(s), collectively called the packaging signal, in the viral RNA [7,8,9,10,11,12,13,14]. In some retroviruses, such as HIV-1, RNA partner selection and the initiation of the dimerization process occur in the cytoplasm [15]; RNA dimerizes before encapsidation [16,17]. As base-pairing is involved in RNA partner selection for the initial dimerization process, the identity of the DIS sequence affects the ability of two HIV-1 RNAs derived from different proviruses to be copackaged together to form heterozygous particles [16]
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