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

Ioana Boeras, Zhenwei Song, Andrew Moran, Jarryd Franklin, William Clay Brown, Marc Johnson, Kathleen Boris-Lawrie, Xiao Heng

Research output: Contribution to journalArticlepeer-review

14 Scopus citations

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.

Original languageEnglish (US)
Pages (from-to)2418-2429
Number of pages12
JournalJournal of Molecular Biology
Volume428
Issue number11
DOIs
StatePublished - Jun 5 2016

Bibliographical note

Funding Information:
This research was supported by the NIH National Institute of General Medical Sciences P50 GM 103297 to K.B.L. and X.H. We are grateful for plasmids: pFL-RHA provided by C.G. Lee (UMDNJ), pMSM-dEnv provided by A.T. Panganiban (TNRPC), LentiCRISPR v2 from Feng Zhang (Addgene plasmid #52961), NL43Rev(-)R(-).S by B. Felber (NCI), and the AIDS Reagent Repository for antisera.

Keywords

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

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