2. Academic Validation
  3. DHX9/RHA Binding to the PBS-Segment of the Genomic RNA during HIV-1 Assembly Bolsters Virion Infectivity

DHX9/RHA Binding to the PBS-Segment of the Genomic RNA during HIV-1 Assembly Bolsters Virion Infectivity

  • J Mol Biol. 2016 Jun 5;428(11):2418-2429. doi: 10.1016/j.jmb.2016.04.011.
Ioana Boeras 1,  Zhenwei Song 2,  Andrew Moran 2,  Jarryd Franklin 2,  William Clay Brown 3,  Marc Johnson 4,  Kathleen Boris-Lawrie 5,  Xiao Heng 6
Affiliations

Affiliations

  • 1 Department of Veterinary and Biomedical Sciences, University of Minnesota, Saint Paul, MN 55108, USA.
  • 2 Department of Biochemistry, University of Missouri, Columbia, MO 65211, USA.
  • 3 Center for Structural Biology, Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109, USA.
  • 4 Department of Molecular Microbiology and Immunology, University of Missouri, Columbia, MO 65211, USA.
  • 5 Department of Veterinary and Biomedical Sciences, University of Minnesota, Saint Paul, MN 55108, USA. Electronic address: [email protected].
  • 6 Department of Biochemistry, University of Missouri, Columbia, MO 65211, USA. Electronic address: [email protected].
PMID: 27107641 DOI: 10.1016/j.jmb.2016.04.011
Abstract

Cellular RNA-binding proteins incorporated into virions during human immunodeficiency virus type 1 (HIV-1) assembly promote the replication efficiency of progeny virions. Despite its critical role in bolstering virion infectivity, the molecular basis for the incorporation of DHX9/RNA helicase A (RHA) to virions remains unclear. Here, cell-based experiments demonstrate that the truncation of segments of the HIV-1 5'-untranslated region (5'-UTR) distinct from the core encapsidation sequence eliminated virion incorporation of RHA, indicating that RHA recruitment is mediated by specific interactions with the HIV-1 5'-UTR. In agreement with biological data, isothermal titration calorimetry determined that the dimer conformation of the 5'-UTR binds one RHA molecule per RNA strand, and the interaction is independent of nucleocapsid protein binding. NMR spectra employing a deuterium-labeling approach enabled resolution of the dimeric 5'-UTR in complex with the RHA N-terminal domain. The structure of the large molecular mass complex was dependent on RHA binding to a double-stranded region of the primer binding site (PBS)-segment of the 5'-UTR. A single A-to-C substitution was sufficient to disrupt biophysical conformation and attenuate virion infectivity in cell-based assays. Taken together, our studies demonstrate the structural basis for HIV-1 genomic RNA to recruit beneficial cellular cofactor to virions. The support of progeny virion infectivity by RHA is attributable to structure-dependent binding at the PBS-segment of the HIV-1 5'-UTR during virus assembly.

Keywords

NMR; PBS-segment; RNA-binding domain; dimeric 5′-UTR; virus infectivity.

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