Rev-dependent association of the intron-containing HIV-1 gag mRNA with thenuclear actin bundles and the inhibition of its nucleocytoplasmic transport by latrunculin-B

Background: A hallmark of HIV-1 gene expression is that unspliced genomic R NA, which also acts as mRNA for the expression of Gag/Pol, is exported to t he cytoplasm. Rev directs this transport through the nuclear export signal (NES). Results: Fluorescence in situ hybridization and immunocytochemistry demonst rated that gag mRNA, Rev, and its NES receptor, CRM1, and RanGTPase formed nuclear tracks which were congruent with underlying beta-actin bundles. Act in bundle formation was confirmed electron-microscopically. These bundles w ere observed upon Rev-containing gag RNP formation. The loss of bundles was associated with the nuclear retention of gag mRNA. Reverse transcription-p olymerase chain reaction analysis of both cytoplasmic and nuclear gag mRNAs demonstrated that disruption of nuclear actin filament formation by latrun culin-B (LAT-B), an F-actin depolymerizing compound, resulted in the dose-d ependent inhibition of gag mRNA export. The differential subtyping of the m RNA-positive cells confirmed morphologically the effect of LAT-B treatment. The export inhibition was specific to gag mRNA and export of fully spliced HIV-1 tat/rev mRNAs as well as cellular GAPDH mRNA was not affected by the compound. Conclusions: Nuclear beta-actin bundles are suggested to be functionally in volved in the Rev-dependent nucleocytoplasmic transport of intron-containin g HIV-1 gag mRNA.