Dissection of specific binding of HIV-1 Gag to the "packaging signal" in viral RNA
Abstract
Selective packaging of HIV-1 genomic RNA (gRNA) requires the presence of a cis-acting RNA element called the “packaging signal” (Ψ). However, the mechanism by which Ψ promotes selective packaging of the gRNA is not well understood. We used fluorescence correlation spectroscopy and quenching data to monitor the binding of recombinant HIV-1 Gag protein to Cy5-tagged 190-base RNAs. At physiological ionic strength, Gag binds with very similar, nanomolar affinities to both Ψ-containing and control RNAs. We challenged these interactions by adding excess competing tRNA; introducing mutations in Gag; or raising the ionic strength. These modifications all revealed high specificity for Ψ. This specificity is evidently obscured in physiological salt by non-specific, predominantly electrostatic interactions. This nonspecific activity was attenuated by mutations in the MA, CA, and NC domains, including CA mutations disrupting Gag-Gag interaction. We propose that gRNA is selectively packaged because binding to Ψ nucleates virion assembly with particular efficiency.
Article and author information
Author details
Funding
National Cancer Institute
- Mauricio Comas-Garcia
- Siddhartha AK Datta
- Alan Rein
Intramural AIDS Targeted Antiviral Therapy Program
- Mauricio Comas-Garcia
- Alan Rein
National Institutes of Health (Contract HHS N26120080001E)
- Prabhakar R Gudla
National Institute of Allergy and Infectious Diseases
- Rajat Varma
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Stephen P Goff, Howard Hughes Medical Institute, Columbia University, United States
Version history
- Received: March 21, 2017
- Accepted: July 13, 2017
- Accepted Manuscript published: July 20, 2017 (version 1)
- Version of Record published: July 27, 2017 (version 2)
- Version of Record updated: July 28, 2017 (version 3)
Copyright
This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.
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