Figure 2. Some Vpr proteins degrade SAMHD1.

(A) Unrooted phylogeny of 115 vpr and vpx sequences among diverse primate lentiviruses. Bootstrap values indicate maximum likelihood proportions that are highly supported by Bayesian inference (Figure S1A). Seven phylogenetic clusters are shaded in colors (cutoff at ML bootstrap >75, Bayesian posterior probability > 0.87). Vpx sequences form 2 clades (Light blue and dark blue shaded) that have strong support of monophyly from all other vpr sequences. Functional phenotype of Vpr and Vpx (Table I) that degrade SAMHD1 (Blue stars) or do not degrade SAMHD1 (Red stars) are overlaid on the phylogeny. See also Figure S1B.

(B) Western blot analysis of Colobus monkey, De Brazza's monkey, and African green monkey (AGM) SAMHD1 in the presence of indicated Vpr constructs. The AGM SAMHD1 tested is from the Vervet subspecies matching the SIVagmVer 9648 host strain, the Colobus SAMHD1 tested is from the Colobus guereza subspecies.

(C) Association of SAMHD1 with Vpr and DDB1 by co-immunoprecipitation was detected by western blot analysis of HA-immunoprecipitated SAMHD1 for FLAG-epitope tagged Vpr and DDB1 association (IP), or input expression (Input). After transfection, cells were treated with 25μM MG-132 for 12 hours prior to immunoprecipitation. SIVdeb Vpr interacts with De Brazza's monkey SAMHD1 (IP), but SAMHD1 expression was not rescued by MG132 treatment (Input). SIVmnd1 Vpr, which fails to degrade mandrill SAMHD1, was assayed as a negative control. Actin and the antibody light chain (Lc) are shown as loading controls.