Expression Level of HIV-1 Vif Can Be Fluctuated by Natural Nucleotide Variations in the vif-Coding and Regulatory SA1D2prox Sequences of the Proviral Genome

Abstract

Vif is required for HIV-1 replication in natural target cells by counteracting host restriction factors, APOBEC3 (A3) proteins. We recently demonstrated that Vif expression level can be changed by naturally occurring single-nucleotide variations within SA1D2prox of the HIV-1 genome. We also found that levels for vif/vpr mRNAs are inversely correlated. While amino acid sequence per se is critical for functionality, Vif expression level modulated by signal sequences in its coding region is likely to be important as well. There are two splicing sites in the region involved in vpr expression. To reveal possible fluctuations of Vif-expression level, we examined SA1D2prox and vif gene by chimeric approaches using HIV-1 subtypes B and C with distinct anti-A3 activity. In this report, recombinant clones in subtype B backbone carrying chimeric sequences with respect to SA1D2prox/vif and those within the vif-coding region were generated. Of these, clones containing vif-coding sequence of subtype C, especially its 3′ region, expressed vif/Vif at a decreased level but did at an increased level for vpr/Vpr. Clones with reduced vif/Vif level grew similarly or slightly better than a parental clone in weakly A3G-positive cells but more poorly in highly A3G-expressing cells. Three clones with this property were also tested for their A3-degrading activity. One of the clones appeared to have some defect in addition to the poor ability to express vif/Vif. Taken all together, our results show that natural variations in the SA1D2prox and vif-coding region can change the Vif-expression level and affect the HIV-1 replication potential.

The Expression Level of HIV-1 Vif Is Optimized by Nucleotide Changes in the Genomic SA1D2prox Region during the Viral Adaptation Process

“…As we have shown, nSNMs within SA1D2prox alter the D1/A1 splicing product levels and concomitantly inversely change the D1/A2 splicing product levels (this study and [ 37 , 38 , 39 ]). In fact, we have confirmed the fluctuation in the Vif expression level for pro-viral clones that carry various chimeric vif sequences containing SA2 and SD3 between subtype B (NL4-3) and subtype C (IndieC) [ 40 ]. Given the long-range interaction between splicing sites [ 62 ], nucleotide sequences that affect the vif mRNA/Vif protein expression level would be present in various areas on the HIV-1 genome.…”

“…We have reported various nSNMs within the SA1D2prox region of the HIV-1 (NL4-3) genome that significantly affect the Vif expression level and viral replication potential ( Figure 1 A) [ 37 , 38 , 39 , 40 ]. Of these mutant clones, NL-D229gat and NL-R224cgc/NL-P238ccg are categorized into low-Vif and excessive-Vif types, respectively ( Figure 1 B).…”

“…Based on comparative sequence analyses of adaptive mutations that emerged in our HIV-1 adaptation system [ 36 ], we have identified a number of naturally-occurring single nucleotide mutations (nSNM) that vary the Vif expression levels within a region around the SA1 and SD2 sites located within the pol -integrase sequence, named the SA1D2prox region ( Figure 1 A). These nSNMs within SA1D2prox that alter the Vif expression levels also affect the replication potential depending on both their Vif expression levels and cellular A3G expression levels [ 37 , 38 , 39 , 40 ]. Pro-viral clones carrying nSNM are grouped into low-, high-, and excessive-Vif types, which express low, high, and extremely high levels of Vif, respectively.…”

Interferon-Regulated Expression of Cellular Splicing Factors Modulates Multiple Levels of HIV-1 Gene Expression and Replication

“…The use of 3′ss A1 can be enhanced or repressed by splicing the regulatory elements that control the recognition of downstream 5′ss D2. It has been shown that the singlenucleotide variations naturally occurring within the SA1D2 proximal region of the HIV-1 genome can modify the Vif expression levels by the alteration of SREs, impacting the inversely correlated levels of vif/vpr mRNAs [237]. We and others demonstrated that Synonymous mutations within viral genomes that lack amino acid alterations can exert an influence on diverse biological processes, including the transcription, RNA structure, splicing, translation, and microRNA targeting.…”

“…The use of 3′ss A1 can be enhanced or repressed by splicing the regulatory elements that control the recognition of downstream 5′ss D2. It has been shown that the single-nucleotide variations naturally occurring within the SA1D2 proximal region of the HIV-1 genome can modify the Vif expression levels by the alteration of SREs, impacting the inversely correlated levels of vif / vpr mRNAs [ 237 ]. We and others demonstrated that infrequently utilized SD2b [ 11 , 238 ] may facilitate vif formation and counteract APOBEC3G [ 218 ].…”