title: Nuclear exportin receptor CAS regulates the NPI-1-mediated nuclear import of HIV-1 Vpr.
abstract: Vpr, an accessory protein of human immunodeficiency virus type 1, is a multifunctional protein that plays an important role in viral replication. We have previously shown that the region between residues 17 and 74 of Vpr (Vpr(N17C74)) contained a bona fide nuclear localization signal and it is targeted Vpr(N17C74) to the nuclear envelope and then imported into the nucleus by importin α (Impα) alone. The interaction between Impα and Vpr is important not only for the nuclear import of Vpr but also for HIV-1 replication in macrophages; however, it was unclear whether full-length Vpr enters the nucleus in a manner similar to Vpr(N17C74). This study investigated the nuclear import of full-length Vpr using the three typical Impα isoforms, Rch1, Qip1 and NPI-1, and revealed that full-length Vpr is selectively imported by NPI-1, but not Rch1 and Qip1, after it makes contact with the perinuclear region in digitonin-permeabilized cells. A binding assay using the three Impα isoforms showed that Vpr bound preferentially to the ninth armadillo repeat (ARM) region (which is also essential for the binding of CAS, the export receptor for Impα) in all three isoforms. Comparison of biochemical binding affinities between Vpr and the Impα isoforms using surface plasmon resonance analysis demonstrated almost identical values for the binding of Vpr to the full-length isoforms and to their C-terminal domains. By contrast, the data showed that, in the presence of CAS, Vpr was released from the Vpr/NPI-1 complex but was not released from Rch1 or Qip1. Finally, the NPI-1-mediated nuclear import of Vpr was greatly reduced in semi-intact CAS knocked-down cells and was recovered by the addition of exogenous CAS. This report is the first to show the requirement for and the regulation of CAS in the functioning of the Vpr-Impα complex.
authors: Viral Infectious Diseases Unit, RIKEN, Hirosawa, Wako, Saitama, Japan.
authors: Eri
authors: E
authors: Takeda
authors: Tomoyuki
authors: T
authors: Murakami
authors: Go
authors: G
authors: Matsuda
authors: Hironobu
authors: H
authors: Murakami
authors: Tamotsu
authors: T
authors: Zako
authors: Mizuo
authors: M
authors: Maeda
authors: Yoko
authors: Y
authors: Aida
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism*; D029563:Cellular Apoptosis Susceptibility Protein / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D004072:Digitonin / Q000494:pharmacology; D055785:Gene Knockdown Techniques; D005982:Glutathione Transferase / Q000378:metabolism; D057809:HEK293 Cells; D006678:HIV; D006367:HeLa Cells; D006801:Humans; D010539:Permeability / Q000187:drug effects; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D020349:Surface Plasmon Resonance; D028901:alpha Karyopherins / Q000737:chemistry / Q000378:metabolism*; D054325:vpr Gene Products, Human Immunodeficiency Virus / Q000378:metabolism*
chemical_list: D029563:Cellular Apoptosis Susceptibility Protein; C423142:KPNA1 protein, human; C106259:KPNA4 protein, human; D020033:Protein Isoforms; D028901:alpha Karyopherins; C087978:karyopherin alpha 2; D054325:vpr Gene Products, Human Immunodeficiency Virus; C514912:vpr protein, Human immunodeficiency virus 1; D005982:Glutathione Transferase; D004072:Digitonin