Limits...
HIV-1 viral infectivity factor (Vif) alters processive single-stranded DNA scanning of the retroviral restriction factor APOBEC3G.

Feng Y, Love RP, Chelico L - J. Biol. Chem. (2013)

Bottom Line: Vif is thought to primarily overcome APOBEC3G through an interaction that mediates APOBEC3G ubiquitination and results in its proteasomal degradation.Specifically, VifHXB2 inhibited the jumping and VifIIIB inhibited the sliding movements of APOBEC3G.That the partially processive APOBEC3G was less effective at inducing mutagenesis in a model HIV-1 replication assay suggests that Vif co-encapsidation with APOBEC3G can promote sublethal mutagenesis of HIV-1 proviral DNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5E5, Canada.

ABSTRACT
APOBEC3G is a retroviral restriction factor that can inhibit the replication of human immunodeficiency virus, type 1 (HIV-1) in the absence of the viral infectivity factor (Vif) protein. Virion-encapsidated APOBEC3G can deaminate cytosine to uracil in viral (-)DNA, which leads to hypermutation and inactivation of the provirus. APOBEC3G catalyzes these deaminations processively on single-stranded DNA using sliding and jumping movements. Vif is thought to primarily overcome APOBEC3G through an interaction that mediates APOBEC3G ubiquitination and results in its proteasomal degradation. However, Vif may also inhibit APOBEC3G mRNA translation, virion encapsidation, and deamination activity. Here we investigated the molecular mechanism of VifIIIB- and VifHXB2-mediated inhibition of APOBEC3G deamination activity. Biochemical assays using a model HIV-1 replication assay and synthetic single-stranded or partially double-stranded DNA substrates demonstrated that APOBEC3G has an altered processive mechanism in the presence of Vif. Specifically, VifHXB2 inhibited the jumping and VifIIIB inhibited the sliding movements of APOBEC3G. The absence of such an effect by Vif on degradation-resistant APOBEC3G D128K indicates that a Vif-APOBEC3G interaction mediates this effect. That the partially processive APOBEC3G was less effective at inducing mutagenesis in a model HIV-1 replication assay suggests that Vif co-encapsidation with APOBEC3G can promote sublethal mutagenesis of HIV-1 proviral DNA.

Show MeSH

Related in: MedlinePlus

VifHXB2 and VifIIIB have a different effect on the decrease in A3G-induced mutations in the prot sequence. Individual analysis of each prot clone enabled determination of the percentage of clones that would result in an unmutated (no mutations; red), mutated and active (black), or mutated and inactive (green) prot. A, A3G was able to inactivate the prot in 60% of clones and left an active prot in 8% of mutated clones. A3G did not induce any mutations in the prot in 32% of clones. B, A3G-induced mutagenesis in the presence of VifHXB2 resulted in fewer mutations overall. There were 68% of clones with no mutations, 24% of clones were mutated and inactive, and 8% of clones were mutated but retained prot activity. C, the effect of VifIIIB on A3G-induced mutagenesis was less than that of VifHXB2 and resulted in 40% of clones with no mutations, 52% of clones with inactive prot, and 8% of clones with mutated but active prot.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3585047&req=5

Figure 2: VifHXB2 and VifIIIB have a different effect on the decrease in A3G-induced mutations in the prot sequence. Individual analysis of each prot clone enabled determination of the percentage of clones that would result in an unmutated (no mutations; red), mutated and active (black), or mutated and inactive (green) prot. A, A3G was able to inactivate the prot in 60% of clones and left an active prot in 8% of mutated clones. A3G did not induce any mutations in the prot in 32% of clones. B, A3G-induced mutagenesis in the presence of VifHXB2 resulted in fewer mutations overall. There were 68% of clones with no mutations, 24% of clones were mutated and inactive, and 8% of clones were mutated but retained prot activity. C, the effect of VifIIIB on A3G-induced mutagenesis was less than that of VifHXB2 and resulted in 40% of clones with no mutations, 52% of clones with inactive prot, and 8% of clones with mutated but active prot.

Mentions: To determine whether each individual model provirus synthesized in our assay would code for an active or inactive protease, we analyzed each clone individually (Fig. 2). A3G-induced mutagenesis was able to cause inactivation of the prot 60% of the time and left the prot active only 8% of the time (Fig. 2A). A significant amount of clones (32%) had no mutations in the prot region. A3G-induced mutagenesis in the presence of VifHXB2 changed this distribution so that 68% of clones had no mutations in the prot and 24% had mutations that inactivated the prot (Fig. 2B). Again, 8% of clones were mutated but still coded for an active prot (Fig. 2B). The same trend was found for A3G in the presence of VifIIIB where slightly more clones were not mutated (40%; Fig. 2C) in comparison with A3G alone (32%; Fig. 2A). These results are in agreement with the analysis of the total population of mutations (supplemental Table S3) where there was an overall decrease in mutations rather than an alteration of the mutational hot spots.


HIV-1 viral infectivity factor (Vif) alters processive single-stranded DNA scanning of the retroviral restriction factor APOBEC3G.

Feng Y, Love RP, Chelico L - J. Biol. Chem. (2013)

VifHXB2 and VifIIIB have a different effect on the decrease in A3G-induced mutations in the prot sequence. Individual analysis of each prot clone enabled determination of the percentage of clones that would result in an unmutated (no mutations; red), mutated and active (black), or mutated and inactive (green) prot. A, A3G was able to inactivate the prot in 60% of clones and left an active prot in 8% of mutated clones. A3G did not induce any mutations in the prot in 32% of clones. B, A3G-induced mutagenesis in the presence of VifHXB2 resulted in fewer mutations overall. There were 68% of clones with no mutations, 24% of clones were mutated and inactive, and 8% of clones were mutated but retained prot activity. C, the effect of VifIIIB on A3G-induced mutagenesis was less than that of VifHXB2 and resulted in 40% of clones with no mutations, 52% of clones with inactive prot, and 8% of clones with mutated but active prot.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3585047&req=5

Figure 2: VifHXB2 and VifIIIB have a different effect on the decrease in A3G-induced mutations in the prot sequence. Individual analysis of each prot clone enabled determination of the percentage of clones that would result in an unmutated (no mutations; red), mutated and active (black), or mutated and inactive (green) prot. A, A3G was able to inactivate the prot in 60% of clones and left an active prot in 8% of mutated clones. A3G did not induce any mutations in the prot in 32% of clones. B, A3G-induced mutagenesis in the presence of VifHXB2 resulted in fewer mutations overall. There were 68% of clones with no mutations, 24% of clones were mutated and inactive, and 8% of clones were mutated but retained prot activity. C, the effect of VifIIIB on A3G-induced mutagenesis was less than that of VifHXB2 and resulted in 40% of clones with no mutations, 52% of clones with inactive prot, and 8% of clones with mutated but active prot.
Mentions: To determine whether each individual model provirus synthesized in our assay would code for an active or inactive protease, we analyzed each clone individually (Fig. 2). A3G-induced mutagenesis was able to cause inactivation of the prot 60% of the time and left the prot active only 8% of the time (Fig. 2A). A significant amount of clones (32%) had no mutations in the prot region. A3G-induced mutagenesis in the presence of VifHXB2 changed this distribution so that 68% of clones had no mutations in the prot and 24% had mutations that inactivated the prot (Fig. 2B). Again, 8% of clones were mutated but still coded for an active prot (Fig. 2B). The same trend was found for A3G in the presence of VifIIIB where slightly more clones were not mutated (40%; Fig. 2C) in comparison with A3G alone (32%; Fig. 2A). These results are in agreement with the analysis of the total population of mutations (supplemental Table S3) where there was an overall decrease in mutations rather than an alteration of the mutational hot spots.

Bottom Line: Vif is thought to primarily overcome APOBEC3G through an interaction that mediates APOBEC3G ubiquitination and results in its proteasomal degradation.Specifically, VifHXB2 inhibited the jumping and VifIIIB inhibited the sliding movements of APOBEC3G.That the partially processive APOBEC3G was less effective at inducing mutagenesis in a model HIV-1 replication assay suggests that Vif co-encapsidation with APOBEC3G can promote sublethal mutagenesis of HIV-1 proviral DNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5E5, Canada.

ABSTRACT
APOBEC3G is a retroviral restriction factor that can inhibit the replication of human immunodeficiency virus, type 1 (HIV-1) in the absence of the viral infectivity factor (Vif) protein. Virion-encapsidated APOBEC3G can deaminate cytosine to uracil in viral (-)DNA, which leads to hypermutation and inactivation of the provirus. APOBEC3G catalyzes these deaminations processively on single-stranded DNA using sliding and jumping movements. Vif is thought to primarily overcome APOBEC3G through an interaction that mediates APOBEC3G ubiquitination and results in its proteasomal degradation. However, Vif may also inhibit APOBEC3G mRNA translation, virion encapsidation, and deamination activity. Here we investigated the molecular mechanism of VifIIIB- and VifHXB2-mediated inhibition of APOBEC3G deamination activity. Biochemical assays using a model HIV-1 replication assay and synthetic single-stranded or partially double-stranded DNA substrates demonstrated that APOBEC3G has an altered processive mechanism in the presence of Vif. Specifically, VifHXB2 inhibited the jumping and VifIIIB inhibited the sliding movements of APOBEC3G. The absence of such an effect by Vif on degradation-resistant APOBEC3G D128K indicates that a Vif-APOBEC3G interaction mediates this effect. That the partially processive APOBEC3G was less effective at inducing mutagenesis in a model HIV-1 replication assay suggests that Vif co-encapsidation with APOBEC3G can promote sublethal mutagenesis of HIV-1 proviral DNA.

Show MeSH
Related in: MedlinePlus