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Mutant human immunodeficiency virus type 1 genomes with defects in RNA dimerization or encapsidation
1Department of Pathology, University of California, San Francisco 94143, USA.
Journal of Virology
|May 1, 1997
Summary
This study reveals that specific RNA structures within the human immunodeficiency virus type 1 (HIV-1) psi locus are crucial for packaging viral RNA and discriminating between spliced and unspliced transcripts. RNA dimerization is important for infectivity but not essential for genomic packaging.
Area of Science:
- Virology
- Molecular Biology
- Structural Biology
Background:
- Retrovirus particles, including human immunodeficiency virus type 1 (HIV-1), contain a dimeric RNA genome.
- The HIV-1 psi locus, located near the 5' end, is critical for RNA dimerization and packaging into virions via interactions with the Gag protein.
- The psi locus is organized into stem-loops, with SL1 implicated in dimer initiation and SL1, SL3, and SL4 containing Gag binding sites.
Purpose of the Study:
- To investigate the roles of specific psi locus RNA structures and Gag binding sites in HIV-1 RNA packaging, dimerization, and infectivity.
- To determine whether RNA dimerization is a prerequisite for genomic packaging or serves an independent function.
Main Methods:
- Creation of HIV-1 proviruses with mutations in the psi locus affecting Gag binding and/or RNA dimerization.
- Characterization of viral assembly and infectivity using a single-step infectious assay.
- Analysis of virion RNA content, including spliced and unspliced transcripts, and assessment of RNA dimerization using nondenaturing gel electrophoresis.
Main Results:
- Mutations eliminating Gag binding sites in SL1 or SL3 caused defects in genomic RNA packaging and infectivity, with increased spliced RNA content in virions.
- The structural integrity of SL1 and SL3 stems, rather than specific sequences, was critical for RNA packaging.
- Disruption of SL1 or its stem impaired genomic dimerization.
- Mutations preventing in vitro dimerization but not affecting virion RNA content led to impaired infectivity.
Conclusions:
- Both SL1 and SL3 contribute to the selective packaging of unspliced genomic RNA over spliced transcripts.
- RNA dimerization is not required for genomic RNA packaging but plays a distinct role in the retroviral infectious cycle.
- The structural conformation of stem-loops within the psi locus is vital for efficient viral RNA packaging and infectivity.