Dissection of specific binding of HIV-1 Gag to the 'packaging signal' in viral RNA

Elife. 2017 Jul 20;6:e27055. doi: 10.7554/eLife.27055.

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.

Keywords: HIV; RNA; biophysics; infectious disease; microbiology; protein-RNA interaction; retrovirus; structural biology; virus; virus assembly.

Publication types

  • Research Support, N.I.H., Intramural
  • Research Support, N.I.H., Extramural

MeSH terms

  • HIV-1 / physiology*
  • Protein Binding
  • RNA, Viral / metabolism*
  • Spectrometry, Fluorescence
  • Virus Assembly*
  • gag Gene Products, Human Immunodeficiency Virus / metabolism*

Substances

  • RNA, Viral
  • gag Gene Products, Human Immunodeficiency Virus