title: Evidence for Vpr-dependent HIV-1 replication in human CD4+ CEM.NKR T-cells.
abstract: BACKGROUND
  Vpr is exclusively expressed in primate lentiviruses and contributes to viral replication and disease progression in vivo. HIV-1 Vpr has two major activities in vitro: arrest of cell cycle in the G2 phase (G2 arrest), and enhancement of viral replication in macrophages. Previously, we reported a potent HIV-1 restriction in the human CD4+ CEM.NKR (NKR) T cells, where wild-type (WT) HIV-1 replication was inhibited by almost 1,000-fold. From the parental NKR cells, we isolated eight clones by limiting dilution. These clones showed three levels of resistance to the WT HIV-1 infection: non-permissive (NP), semi-permissive (SP), and permissive (P). Here, we compared the replication of WT, Vif-defective, Vpr-defective, and Vpu-defective viruses in these cells.
  
  
  RESULTS
  Although both WT and Vpu-defective viruses could replicate in the permissive and semi-permissive clones, the replication of Vif-defective and Vpr-defective viruses was completely restricted. The expression of APOBEC3G (A3G) cytidine deaminase in NKR cells explains why Vif, but not Vpr, was required for HIV-1 replication. When the Vpr-defective virus life cycle was compared with the WT virus life cycle in the semi-permissive cells, it was found that the Vpr-defective virus could enter the cell and produce virions containing properly processed Gag and Env proteins, but these virions showed much less efficiency for reverse transcription during the next-round of infection. In addition, although viral replication was restricted in the non-permissive cells, treatment with arsenic trioxide (As2O3) could completely restore WT, but not Vpr-defective virus replication. Moreover, disruption of Vpr binding to its cofactor DCAF1 and/or induction of G2 arrest activity did not disrupt the Vpr activity in enhancing HIV-1 replication in NKR cells.
  
  
  CONCLUSIONS
  These results demonstrate that HIV-1 replication in NKR cells is Vpr-dependent. Vpr promotes HIV-1 replication from the 2nd cycle likely by overcoming a block at early stage of viral replication; and this activity does not require DCAF1 and G2 arrest. Further studies of this mechanism should provide new understanding of Vpr function in the HIV-1 life cycle.
authors.affiliation: Department of Microbiology and Molecular Genetics, Michigan State University, 567 Wilson Road, 2215, Biomedical and Physical Sciences Building, East Lansing, MI 48824-4320, USA.
authors.forename: Tao
authors.initials: T
authors.lastname: Zhou
authors.forename: Ying
authors.initials: Y
authors.lastname: Dang
authors.forename: Jacob J
authors.initials: JJ
authors.lastname: Baker
authors.forename: Jiajun
authors.initials: J
authors.lastname: Zhou
authors.forename: Yong-Hui
authors.initials: YH
authors.lastname: Zheng
mesh_terms: D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism* / Q000821:virology*; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D059565:G2 Phase Cell Cycle Checkpoints; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D009154:Mutation; D017346:Protein Serine-Threonine Kinases; D044767:Ubiquitin-Protein Ligases; D056189:Viral Tropism; D014779:Virus Replication; D054325:vpr Gene Products, Human Immunodeficiency Virus / Q000235:genetics / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D054325:vpr Gene Products, Human Immunodeficiency Virus; D044767:Ubiquitin-Protein Ligases; C530533:DCAF1 protein, human; D017346:Protein Serine-Threonine Kinases