Lentiviral Vif degrades the APOBEC3Z3/APOBEC3H protein of its mammalian host and is capable of cross-species activity

Abstract

All lentiviruses except equine infectious anemia virus (EIAV) use the small accessory protein Vif to counteract the restriction activity of the relevant APOBEC3 (A3) proteins of their host species. Prior studies have suggested that the Vif-A3 interaction is species specific. Here, using the APOBEC3H (Z3)-type proteins from five distinct mammals, we report that this is generally not the case: some lentiviral Vif proteins are capable of triggering the degradation of both the A3Z3-type protein of their normal host species and those of several other mammals. For instance, SIV(mac) Vif can mediate the degradation of the human, macaque, and cow A3Z3-type proteins but not of the sheep or cat A3Z3-type proteins. Maedi-visna virus (MVV) Vif is similarly promiscuous, degrading not only sheep A3Z3 but also the A3Z3-type proteins of humans, macaques, cows, and cats. In contrast to the neutralization capacity of these Vif proteins, human immunodeficiency virus (HIV), bovine immunodeficiency virus (BIV), and feline immunodeficiency virus (FIV) Vif appear specific to the A3Z3-type protein of their hosts. We conclude, first, that the Vif-A3Z3 interaction can be promiscuous and, second, despite this tendency, that each lentiviral Vif protein is optimized to degrade the A3Z3 protein of its mammalian host. Our results thereby suggest that the Vif-A3Z3 interaction is relevant to lentivirus biology.

FIG. 1.

Comparison of A3Z3 and Vif sequences. (A) The relatedness of the mammals relevant to this study. The numbers at the branch nodes are approximate divergence times in millions of years. (Adapted from reference 22). (B) Percent sequence identity and similarity between all the mammalian A3Z3 proteins. Abbreviations for different mammals: Hs, human; Rh, rhesus macaque; Bt, cow; Oa, sheep; Fc, cat. (C) Alignment of the ELOC binding motifs of the lentiviral Vifs used in this study. The numbers are the first and last residue numbers.

FIG. 2.

Antiretroviral properties of cow A3 proteins and their sensitivity to BIV Vif. (A) Relative infectivity of HIV-GFP produced in the presence of the indicated A3s and a vector control (open bars), BIV Vif (black bars), or BIV Vif_SLQ-AAA (gray bars). The double vector control (empty pcDNA3.1 and pVR1012 vectors) was normalized to 1, and the error bars report the standard errors of the means from 3 independent experiments. (B) Immunoblots showing the producer cell levels of A3 proteins (anti-HA), Vif (anti-myc), and tubulin. −, vector control; +, wild-type Vif; m, Vif_SLQ-AAA mutant.

FIG. 3.

Sensitivity of mammalian A3Z3 proteins to BIV and MVV Vif. (A) Relative infectivity of HIV-GFP produced in the presence of the indicated A3s and a vector control (open bars), BIV Vif (black bars), or BIV Vif_SLQ-AAA (gray bars). The double vector control was normalized to 1, and the error bars report the standard errors of the means from 3 independent experiments. Producer cell immunoblots are shown below for A3 proteins (anti-HA), Vif (anti-myc), and tubulin. The mouse A3 signals were faint on the original full immunoblot, and so an image of a longer exposure was inserted in the identical position for presentation. (B) Relative infectivity of HIV-GFP produced in the presence of the indicated A3s and a vector control (open bars), MVV Vif (black bars), or MVV Vif_SLQ-AAA (gray bars). The figure is organized and labeled as described above.

FIG. 4.

Sensitivity of mammalian A3Z3 proteins to HIV-1_LAI Vif. (A) Relative infectivity of HIV-GFP produced in the presence of the indicated A3s and a vector control (open bars), HIV-1_LAI Vif (black bars), or HIV-1_LAI Vif_SLQ-AAA (gray bars). The double vector control was normalized to 1, and the error bars report the standard errors of the means from 3 independent experiments. Producer cell immunoblots are shown below for A3 proteins (anti-HA), Vif (anti-myc), and tubulin. (B) The relative infectivity of HIV-GFP produced in the presence of untagged human A3H (haplotype II) or A3H-HA and a vector control (open bars), HIV-1_LAI Vif (black bars), or HIV-1_LAI Vif_SLQ-AAA (gray bars). Producer cell immunoblots are shown below for A3H (anti-A3H), Vif (anti-myc), and tubulin.

FIG. 5.

Sensitivity of mammalian A3Z3 proteins to SIV_mac and FIV Vif. (A) The relative infectivity of HIV-GFP produced in the presence of the indicated A3s and a vector control (open bars), SIV_macVif (black bars), or SIV_mac Vif_SLQ-AAA (gray bars). The double vector control was normalized to 1, and the error bars report the standard errors of the means from 3 independent experiments. Producer cell immunoblots are shown below for A3 proteins (anti-HA), Vif (anti-myc), and tubulin. (B) Relative infectivity of HIV-GFP produced in the presence of the indicated A3s and a vector control (open bars), FIV Vif (black bars), or FIV Vif_TLQ-AAA (gray bars). The figure is labeled as described above. We note that the transfer of protein on this particular blot is slightly less efficient toward the right, but this does not alter our overall conclusions (Table 1).