Quantitative mass spectrometry of DENV-2 RNA-interacting proteins reveals that the DEAD-box RNA helicase DDX6 binds the DB1 and DB2 3’ UTR structures

Dengue virus (DENV) is a rapidly re-emerging flavivirus that causes dengue fever (DF), dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS), diseases for which there are no available therapies or vaccines.  The DENV-2 positive-strand RNA genome contains 5’ and 3’ untranslated regions (UTRs) that have been shown to form secondary structures required for virus replication and interaction with host cell proteins.  In order to comprehensively identify host cell factors that bind the DENV-2 UTRs, we performed RNA chromatography, using the DENV-2 5’ and 3’ UTRs as “bait”, combined with quantitative mass spectrometry.  We identified several proteins, including DDX6, G3BP1, G3BP2, Caprin1, and USP10, implicated in P body (PB) and stress granule (SG) function, and not previously known to bind DENV RNAs.  Indirect immunofluorescence microscopy showed these proteins to colocalize with the DENV replication complex.  Moreover, DDX6 knockdown resulted in reduced amounts of infectious particles and viral RNA i...