ARGININE RESIDUES IN THE C-TERMINUS OF HIV-1 VPR ARE IMPORTANT FOR NUCLEAR-LOCALIZATION AND CELL-CYCLE ARREST

HIV-1 viral protein R (Vpr) is predominantly localized to the nucleus and plays an important role for viral preintegration complex import in to the nucleus. In this study, we investigated the influence on subcel lular localization of Arg residues in the C-terminus of Vpr. Consisten t with previous studies, about 90% of the cells manifested diffuse nuc lear staining in the Vpr-expressed cells. Besides diffuse nuclear stai ning, punctate perinuclear staining, and punctate cytoplasmic staining were also observed in the immunofluorescence studies. Deletion of the Ser-Arg-lle-Gly residues (amino acids 79-82; SRIG) had no effect on t he Vpr localization. However, deletion of the Arg-Gln-Arg-Arg residues (amino acids 85-88; RQRR) resulted in a smooth perinuclear staining p attern. Substitution of five Arg residues with Asn (amino acids 80, 85 , 87, 88, and 90; R --> N5) resulted in a diffuse cytoplasmic staining . Subcellular fractionation analyses support the immunofluorescence st aining results. These findings indicate that the C-terminal Arg residu es of HIV-1 Vpr play an important role for Vpr nuclear localization. A ll the Vpr mutants were appropriately expressed, exhibited no signific ant defect on the protein stability, and were incorporated efficiently into virus-like particles. Both SRIG and R --> N5 mutants lost their cell cycle arrest activities and the RQRR deletion only exhibited a lo w level of cell arrest activity. Therefore, the Arg residues in the HI V-1 Vpr C-terminus are important for Vpr nuclear localization and cell cycle arrest, but had no effect on protein stability or Vpr incorpora tion into virus-like particles. (C) 1998 Academic Press.