Distinct Determinants in HIV-1 Vif and Human APOBEC3 Proteins Are Required for the Suppression of Diverse Host Anti-Viral Proteins

Abstract

BackgroundAPOBEC3G (A3G) and related cytidine deaminases of the APOBEC3 family of proteins are potent inhibitors of many retroviruses, including HIV-1. Formation of infectious HIV-1 requires the suppression of multiple cytidine deaminases by Vif. HIV-1 Vif suppresses various APOBEC3 proteins through the common mechanism of recruiting the Cullin5-ElonginB-ElonginC E3 ubiquitin ligase to induce target protein polyubiquitination and proteasome-mediated degradation. The domains in Vif and various APOBEC3 proteins required for APOBEC3 recognition and degradation have not been fully characterized.Methods and FindingsIn the present study, we have demonstrated that the regions of APOBEC3F (A3F) that are required for its HIV-1-mediated binding and degradation are distinct from those reported for A3G. We found that the C-terminal cytidine deaminase domain (C-CDD) of A3F alone is sufficient for its interaction with HIV-1 Vif and its Vif-mediated degradation. We also observed that the domains of HIV-1 Vif that are uniquely required for its functional interaction with full-length A3F are also required for the degradation of the C-CDD of A3F; in contrast, those Vif domains that are uniquely required for functional interaction with A3G are not required for the degradation of the C-CDD of A3F. Interestingly, the HIV-1 Vif domains required for the degradation of A3F are also required for the degradation of A3C and A3DE. On the other hand, the Vif domains uniquely required for the degradation of A3G are dispensable for the degradation of cytidine deaminases A3C and A3DE.ConclusionsOur data suggest that distinct regions of A3F and A3G are targeted by HIV-1 Vif molecules. However, HIV-1 Vif suppresses A3F, A3C, and A3DE through similar recognition determinants, which are conserved among Vif molecules from diverse HIV-1 strains. Mapping these determinants may be useful for the design of novel anti-HIV inhibitors.

A Single Amino Acid Difference in Human APOBEC3H Variants Determines HIV-1 Vif Sensitivity

“…293T and MAGI-CCR5 (AIDS Research Reagents Program) cells were maintained in Dulbecco's modified Eagle's medium (Invitrogen) with 10% fetal bovine serum and 25 g/ml gentamicin (Sigma) and transfected or infected as previously described (92). The following antibodies were used as previously described (96): anti-HA monoclonal antibody (MAb) (Covance, MMS-101R-10000), anti-myc MAb (Sigma, M5546), and antiactin MAb (Sigma, A3853). The anti-Vif antibody was obtained from the AIDS Research Reagents Program (2221), mouse anti-V5 antibody was from Invitrogen (R96025), rabbit anti-V5 was from Covance (PRB-198P), and rabbit anti-green fluorescent protein (anti-GFP) antibody was from Abcam (ab6556-25).…”

“…The eluted materials were then analyzed by SDS-PAGE and immunoblotting with the appropriate antibodies. The anti-HA antibody-agarose conjugate and anti-HA antibody used have been previously described (96).…”

“…Homology modeling of the N-terminal domain of A3G (A3G-NTD) and A3H_HapII was carried out using both the program Modeler (66) and the automated modeling server I-TASSER (96), which has been ranked as the best server in the recent CASP7 and CASP8 modeling contests (1). Modeler and I-TASSER produced very similar models, suggesting convincing modeling results.…”

“…For example, several positively charged amino acids from position 22 to 44 are important for Vif-mediated suppression of A3G but not of A3F (9,21,53,64,74,87,96). In contrast, amino acids 11 to 17 and 74 to 79 of Vif are crucial for the Vif-mediated suppression of A3F but not A3G (32,64,68,74,80).…”

Possible Footprints of APOBEC3F and/or Other APOBEC3 Deaminases, but Not APOBEC3G, on HIV-1 from Patients with Acute/Early and Chronic Infections

“…Both overlapping and distinct Vif domains are involved in the binding of APOBEC3F and APOBEC3G and other deaminases, such as, e.g., APOBEC3C and APOBEC3DE (10,40,56,(71)(72)(73)(74)(75)(76); however, it is likely that a single Vif molecule binds to only one APOBEC3 molecule. Because the number of Vif molecules in HIV-infected cells at any given time point is finite, we hypothesize that viral selection may favor the preferential binding of Vif to APOBEC3G because it would confer a greater survival advantage to HIV-1 than binding to other deaminases.…”