Inhibition of human immunodeficiency virus type 1 Rev function by a dominant-negative mutant of Sam68 through sequestration of unspliced RNA at perinuclear bundles.
Human immunodeficiency virus (HIV) type 1 encodes an essential protein, Rev, which functions to transport unspliced and singly spliced viral transcripts from the nucleus to the cytoplasm to allow expression of the viral structural proteins. It has previously been reported that Sam68 synergistically stimulates Rev activity (T. Reddy et al., Nat. Med. 5:635-642, 1999). Here we report that the Sam68-like mammalian proteins SLM1 and SLM2 also stimulate Rev activity. Their stimulation ability cannot be attributed to a shuttling property, since Sam68, SLM1, and SLM2 do not display significant shuttling activity alone or in the presence of Rev. In addition, Sam68, SLM1, and SLM2 do not affect the equilibrium between unspliced and completely spliced HIV RNA. The C-terminally truncated Sam68 mutant (Sam68DeltaC) previously observed to inhibit the Sam68-mediated stimulation of Rev activity (Reddy et al., 1999) also inhibits SLM1- and SLM2-mediated stimulation of Rev activity. This suggests that the mechanism by which Sam68, SLM1, and SLM2 stimulate Rev activity may be common. Sam68DeltaC does not inhibit Rev activity by inhibiting Rev from shuttling between the nucleus and cytoplasm. Inhibition by Sam68DeltaC is a consequence of its mislocalization to the cytoplasm, as evidenced by the fact that addition of an exogenous nuclear localization signal to Sam68DeltaC restores nuclear localization and stimulation of Rev activity. We demonstrate that Sam68DeltaC causes perinuclear accumulation of unspliced HIV env RNA and propose that Sam68DeltaC inhibits Rev activity by sequestering Rev-responsive RNA away from the translation apparatus.
Soros
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Department of Medical and Molecular Genetics and Microbiology, University of Toronto, Toronto, Ontario M5S 1A8.
Carvajal
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HV
Richard
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Cochrane
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D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D048868:Adaptor Proteins, Signal Transducing; D004268:DNA-Binding Proteins; D015967:Gene Expression Regulation, Viral; D015694:Gene Products, rev / Q000378:metabolism*; D005799:Genes, Dominant; D015497:HIV-1; D006367:HeLa Cells; D006801:Humans; D009154:Mutation; D019913:Nuclear Localization Signals; D012326:RNA Splicing; D012367:RNA, Viral / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000494:pharmacology / Q000502:physiology*; D014162:Transfection; D054321:rev Gene Products, Human Immunodeficiency Virus
D048868:Adaptor Proteins, Signal Transducing; D004268:DNA-Binding Proteins; D015694:Gene Products, rev; C489559:KHDRBS1 protein, human; C485572:KHDRBS2 protein, human; C118777:KHDRBS3 protein, human; C513253:Khdrbs1 protein, mouse; C486528:Khdrbs3 protein, mouse; D019913:Nuclear Localization Signals; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D054321:rev Gene Products, Human Immunodeficiency Virus