Identification of host proteins differentially associated with HIV-1 RNA splice variants

• Main Authors: Rachel Knoener, Edward Evans III, Jordan T Becker, Mark Scalf, Bayleigh Benner, Nathan M Sherer, Lloyd M Smith
• Format: Article
• Language: English
• Published: eLife Sciences Publications Ltd 2021-02-01
• Series: eLife
• Subjects: HIV; splicing; proteomics; RNA binding proteins; RNA imaging; interactome
• Online Access: https://elifesciences.org/articles/62470

HIV-1 generates unspliced (US), partially spliced (PS), and completely spliced (CS) classes of RNAs, each playing distinct roles in viral replication. Elucidating their host protein ‘interactomes’ is crucial to understanding virus-host interplay. Here, we present HyPR-MSSV for isolation of US, PS, and CS transcripts from a single population of infected CD4+ T-cells and mass spectrometric identification of their in vivo protein interactomes. Analysis revealed 212 proteins differentially associated with the unique RNA classes, including preferential association of regulators of RNA stability with US and PS transcripts and, unexpectedly, mitochondria-linked proteins with US transcripts. Remarkably, >80 of these factors screened by siRNA knockdown impacted HIV-1 gene expression. Fluorescence microscopy confirmed several to co-localize with HIV-1 US RNA and exhibit changes in abundance and/or localization over the course of infection. This study validates HyPR-MSSV for discovery of viral splice variant protein interactomes and provides an unprecedented resource of factors and pathways likely important to HIV-1 replication.