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J Virol. 2019 Aug 14. pii: JVI.01048-19. doi: 10.1128/JVI.01048-19. [Epub ahead of print]

Genome wide analysis of hnRNP binding to HIV-1 RNA reveals a key role for hnRNP H1 in alternative viral mRNA splicing.

Author information

1
Department of Molecular Microbiology, Washington University School of Medicine, Saint Louis, MO 63110, USA pbieniasz@rockefeller.edu kutluay@wustl.edu.
2
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
3
Department of Chemistry, Case Western Reserve University, Cleveland, OH 44106, USA.
4
Department of Molecular Microbiology, Washington University School of Medicine, Saint Louis, MO 63110, USA.
5
Laboratory of Retrovirology the Rockefeller University, New York, NY 10028, USA.
6
Department of Biochemistry and Biophysics, and UNC Center For AIDS Research, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
7
Laboratory of Retrovirology the Rockefeller University, New York, NY 10028, USA pbieniasz@rockefeller.edu kutluay@wustl.edu.
8
Howard Hughes Medical Institute, the Rockefeller University, New York, NY 10028, USA.

Abstract

Alternative splicing of HIV-1 mRNAs increases viral coding potential and controls the levels and timing of gene expression. HIV-1 splicing is regulated in part by heterogeneous nuclear ribonucleoproteins (hnRNPs) and their viral target sequences, that typically repress splicing when studied outside their native viral context. Here, we determined the location and extent of hnRNP binding to HIV-1 mRNAs and their impact on splicing in a native viral context. Notably, hnRNPA1, hnRNPA2 and hnRNPB1 bound to many dispersed sites across viral mRNAs. Conversely, hnRNPH1 bound to a few discrete purine-rich sequences, a finding that was mirrored in vitro HnRNPH1 depletion and mutation of a prominent viral RNA hnRNPH1 binding site decreased use of splice acceptor A1, causing a deficit in Vif expression and replicative fitness. This quantitative framework for determining the regulatory inputs governing alternative HIV-1 splicing revealed an unexpected splicing enhancer role for hnRNPH1 through binding to its target element.IMPORTANCE Alternative splicing of HIV-1 mRNAs is an essential, yet quite poorly understood step of virus replication that enhances the coding potential of the viral genome and allows temporal regulation of viral gene expression. Although HIV-1 constitutes an important model system for general studies of the regulation of alternative splicing, the inputs that determine the efficiency with which splice sites are utilized remain poorly defined. Our studies provide an experimental framework to study an essential step of HIV-1 replication more comprehensively and in much greater detail than was previously possible and reveal novel cis acting elements regulating HIV-1 splicing.

PMID:
31413137
DOI:
10.1128/JVI.01048-19

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