Human immunodeficiency virus-1 (HIV-1) Vif is essential for viral evasion of host antiviral factor CEM15/APOBEC3G. We report that Vif interacts with cellular proteins Cul5, elongins B and C, and Rbx1 to form an Skp1-cullin-F-box (SCF)-like complex. The ability of Vif to suppress antiviral activity of APOBEC3G was specifically dependent on Cul5-SCF function, allowing Vif to interact with APOBEC3G and induce its ubiquitination and degradation. A Vif mutant that interacted with APOBEC3G but not with Cul5-SCF was functionally inactive. The Cul5-SCF was also required for Vif function in distantly related simian immunodeficiency virus mac. These results indicate that the conserved Cul5-SCF pathway used by Vif is a potential target for antiviral development.

APOBEC3G, which induces hypermutations in newly synthesized viral DNA, is suppressed by HIV-1 Vif, acting through Cul5-ElonginB-ElonginC E3 ubiquitin ligase. We have now characterized a novel SOCS box in HIV-1 Vif that mediates its interaction with ElonginC. In this SOCS box, alanine replaces the consensus cysteine in the previously identified SOCS box. This new motif was necessary but insufficient for interaction with Cul5-ElonginB-ElonginC, as two highly conserved Cys residues outside the SOCS box were required to interact with Cul5 but not ElonginC. Therefore, selective assembly with Cul5 versus Cul2 E3 may require protein interfaces besides the SOCS-box-ElonginC interaction. Received August 16, 2004; revised version accepted September 24, 2004. APOBEC3G mediates antiviral activity by inducing hypermutations in newly synthesized viral minus-strand DNA (Sheehy et al.

APOBEC3G exerts its antiviral activity by targeting to retroviral particles and inducing viral DNA hypermutations in the absence of Vif. However, the mechanism by which APOBEC3G is packaged into virions remains unclear. We now report that viral genomic RNA enhances but is not essential for human APOBEC3G packaging into human immunodeficiency virus type 1 (HIV-1) virions. Packaging of APOBEC3G was also detected in HIV-1 Gag virus-like particles (VLP) that lacked all the viral genomic RNA packaging signals. Human APOBEC3G could be packaged efficiently into a divergent subtype HIV-1, as well as simian immunodeficiency virus, strain mac, and murine leukemia virus Gag VLP. Cosedimentation of human APOBEC3G and intracellular Gag complexes was detected by equilibrium density and velocity sucrose gradient analysis. Interaction between human APOBEC3G and HIV-1 Gag was also detected by coimmunoprecipitation experiments. This interaction did not require p6, p1, or the C-terminal region of NCp7. However, the N-terminal region, especially the first 11 amino acids, of HIV-1 NCp7 was critical for HIV-1 Gag and APOBEC3G interaction and virion packaging. The linker region flanked by the two active sites of human APOBEC3G was also important for efficient packaging into HIV-1 Gag VLP. Association of human APOBEC3G with RNA-containing intracellular complexes was observed. These results suggest that the N-terminal region of HIV-1 NC, which is critical for binding to RNA and mediating Gag-Gag oligomerization, plays an important role in APOBEC3G binding and virion packaging.The Vif protein, which modulates viral infectivity (15, 21-23, 29, 38, 45, 59, 62, 78-80, 82, 84, 88) and pathogenicity (13, 28, 29, 33, 34,52), is present in nearly all lentiviruses, excluding equine infectious anemia virus. Recently, it has been shown that APOBEC3G has antiviral activity and that its activity is suppressed by Vif (73). APOBEC3G belongs to a family of proteins that have cytidine deaminase activity (35,73,89), and its cellular function is still unknown (35,73,89). Vif mutant, but not wild-type, human immunodeficiency virus type 1 (HIV-1) produced in the presence of APOBEC3G undergoes hypermutations in newly synthesized viral DNA (30, 44,53,55,94), presumably due to C-to-U modification during minus-strand viral DNA synthesis (30, 44,53,55,94).HIV-1 Vif can be coimmunoprecipitated with human APOBEC3G from virus-infected or -transfected cells, suggesting that Vif and APOBEC3G may form a complex (39,56,74,81,93). However, whether Vif directly interacts with APOBEC3G is still unclear (57). Amino acids 54 to 124 of human APOBEC3G have been shown to mediate interaction with HIV-1 Vif (9). Recent observations indicate that a single amino acid variation (at position 128) between human and monkey APOBEC3G determines the species-specific sensitivity to Vif of primate immunodeficiency viruses (4, 54, 71). However, whether this position is involved in APOBEC3G interaction with Vif (4, 54, 71, 91) or whether it is involved in direct interaction between these ...