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Concept: Viral infectivity factor


The human apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3, referred to as A3) proteins are cellular cytidine deaminases that potently restrict retrovirus replication. However, HIV-1 viral infectivity factor (Vif) counteracts the antiviral activity of most A3 proteins by targeting them for proteasomal degradation. To date, the structure of an A3 protein containing a Vif-binding interface has not been solved. Here, we report a high-resolution crystal structure of APOBEC3C and identify the HIV-1 Vif-interaction interface. Extensive structure-guided mutagenesis revealed the role of a shallow cavity composed of hydrophobic or negatively charged residues between the α2 and α3 helices. This region is distant from the DPD motif (residues 128-130) of APOBEC3G that participates in HIV-1 Vif interaction. These findings provide insight into Vif-A3 interactions and could lead to the development of new pharmacologic anti-HIV-1 compounds.

Concepts: HIV, Protein, Cell, Metabolism, Enzyme, Interaction, Viral infectivity factor, APOBEC3G


Human APOBEC3 cytidine deaminases are intrinsic resistance factors to HIV-1. However, HIV-1 encodes a viral infectivity factor (Vif) that degrades APOBEC3 proteins. In vitro APOBEC3F (A3F) anti-HIV-1 activity is weaker than A3G but is partially resistant to Vif degradation unlike A3G. It is unknown whether A3F protein affects HIV-1 disease in vivo. To assess the effect of A3F gene on host susceptibility to HIV- acquisition and disease progression, we performed a genetic association study in six well-characterized HIV-1 natural cohorts. A common six-Single Nucleotide Polymorphism (SNP) haplotype of A3F tagged by a codon-changing variant (p. I231V, with allele (V) frequency of 48% in European Americans) was associated with significantly lower set-point viral load and slower rate of progression to AIDS (Relative Hazards (RH) = 0.71, 95% CI: 0.56, 0.91) and delayed development of pneumocystis pneumonia (PCP) (RH = 0.53, 95% CI: 0.37-0.76). A validation study in the International Collaboration for the Genomics of HIV (ICGH) showed a consistent association with lower set-point viral load. An in vitro assay revealed that the A3F I231V variant may influence Vif mediated A3F degradation. Our results provide genetic epidemiological evidence that A3F modulates HIV-1/AIDS disease progression.

Concepts: HIV, AIDS, Gene, Genetics, Epidemiology, RNA, Viral infectivity factor, APOBEC3G


The crystal structure of viral infectivity factor (Vif) was reported recently, which makes it possible to design new inhibitors against Vif by structure-based drug design. Through analysis of the protein surface of Vif, the C2 pocket located in the N-terminal was found, which is suit for developing small molecular inhibitors. Then, in our article, fragment-based virtual screening (FBVS) was conducted and a series of fragments was obtained, among which, Zif-1 bearing indole scaffold and pyridine ring can form H-bonds with Tyr148 and Ile155. Subsequently, 19 derivatives of Zif-1 were synthesized. Through the immune-fluorescence staining and Western blot assays, Zif-15 shows potent activity in inhibiting Vif-mediated A3G degradation. Further docking experiment shows that Zif-15 form H-bond interactions with residues His139, Tyr148 and Ile155. Therefore, Zif-15 is a promising lead compound against Vif that can be used to treat AIDS.

Concepts: Pharmacology, Bioinformatics, Molecular biology, Drug discovery, Drug design, Virtual screening, Medicinal chemistry, Viral infectivity factor


Lentiviruses threaten human and animal health. Virion infectivity factor (Vif) is essential for the infectivity of most lentiviruses, except for the equine infectious anaemia virus (EIAV). Vif promotes viral infectivity by recruiting a Cullin-based E3 ligase to induce the degradation of a class of host restriction factors, named APOBEC3. Core binding factor beta (CBF-β) is necessary for several primate lentiviral Vif functions, including HIV-1 Vif. Although much progress has been made in understanding the contribution of CBF-β to Vif function, the precise mechanism has not yet been fully elucidated. In this study, we found that an interaction with CBF-β altered the oligomerization and subcellular distribution pattern and increased the stability of two primate lentiviral Vifs, HIV-1 Vif and Macaca simian immunodeficiency virus (SIVmac) Vif. Moreover, using a CBF-β loss-of-function mutant, we demonstrated that the interaction between CBF-β and Vif was not sufficient for Vif assistance; a region including F68 in CBF-β was also required for the stability and function of Vif. For the first time, this study separates the binding and regulating processes of CBF-β when it is promoting Vif function, which further extends our understanding of the biochemical regulation of Vif by CBF-β.

Concepts: HIV, DNA, Protein, RNA interference, Primate, Lentivirus, Viral infectivity factor, Simian immunodeficiency virus


Apolipoprotein B mRNA editing enzyme catalytic polypeptide-like 3 (APOBEC3) proteins are mammalian-specific cellular deaminases and have a robust ability to restrain lentivirus replication. To antagonize APOBEC3-mediated antiviral action, lentiviruses have acquired viral infectivity factor (Vif) as an accessory gene. Mammalian APOBEC3 proteins inhibit lentiviral replication by enzymatically inserting G-to-A hypermutations in the viral genome, whereas lentiviral Vif proteins degrade host APOBEC3 via the ubiquitin/proteasome-dependent pathway. Recent investigations provide evidence that lentiviral vif genes evolved to combat mammalian APOBEC3 proteins. In corollary, mammalian APOBEC3 genes are under Darwinian selective pressure to escape from antagonism by Vif. Based on these observations, it is widely accepted that lentiviral Vif and mammalian APOBEC3 have co-evolved and this concept is called an “evolutionary arms race.” This review provides a comprehensive summary of current knowledge with respect to the evolutionary dynamics occurring at this pivotal host-virus interface.

Concepts: DNA, Protein, Gene, Genetics, Evolution, Virus, RNA, Viral infectivity factor


Viral infectivity factor (Vif) is protective against APOBEC3G (A3G)-mediated viral cDNA hypermutations, and development of molecules that inhibit Vif mediated A3G degradation is a novel strategy for blocking HIV-1 replication. Through optimizations of the central ring of N-(2-methoxyphenyl)-2-((4-nitrophenyl)thio)benzamide (RN-18), we found a potent compound 12c with EC50 value of 1.54 μM, enhancing the antiviral activity more than 150-fold compared with RN-18 in nonpermissive H9 cells. 12c protected A3G from degradation by inhibiting Vif function. Besides, 12c suppressed different HIV-1 clinical strains (HIV-1KM018, HIV-1TC-1 and HIV-1WAN) and drug-resistant strains (NRTI, NNRTI, PI, and FI) with relatively high activities. Amidation of 12c with glycine gave a prodrug 13a, improving the water solubility about 2600-fold compared with 12c. Moreover, 13a inhibited the virus replication efficiently with an EC50 value of 0.228 μM. These results suggested that the prodrug 13a is a promising candidate agent for the treatment of AIDS.

Concepts: Antiretroviral drug, DNA, Pharmacology, Virus, Molecule, Enzyme inhibitor, Viral infectivity factor, APOBEC3G


The retroviral restriction factors of the APOBEC3 (A3) cytidine deaminase family catalyze the deamination of cytidines in single-stranded viral DNA. APOBEC3C (A3C) is a strong antiviral factor against viral infectivity factor (vif)-deficient simian immunodeficiency (SIV) Δvif, however, a weak inhibitor against human immunodeficiency virus (HIV)-1 for reasons unknown. The precise link between the antiretroviral effect of A3C and its catalytic activity is incompletely understood. Here we show that the S61P mutation in human A3C (A3C.S61P) boosted hypermutation in the viral genomes of SIVΔvif and murine leukemia virus but not in human immunodeficiency virus HIV-1Δvif. The enhanced antiviral activity of A3C.S61P correlated with enhanced in vitro cytidine deamination. Furthermore, the S61P mutation did not change substrate specificity of A3C, ribonucleoprotein complex formation, self-association, Zinc coordination or viral incorporation features. We propose that local structural changes induced by the serine-to-proline substitution are responsible for the gain of catalytic activity of A3C.S61P. Our results are a first step towards an understanding of A3C’s DNA binding capacity, deamination-dependent editing, and antiviral functions at the molecular level. We conclude that enhanced enzymatic activity of A3C is insufficient to restrict HIV-1, indicating an unknown escape mechanism of HIV-1.

Concepts: HIV, DNA, Ultraviolet, Metabolism, Enzyme, Genome, Catalysis, Viral infectivity factor


Open reading frame virion infectivity factor (Vif) is conserved among most lentiviruses. Vif molecules contribute to viral replication by inactivating host anti-viral factors, the APOBEC3 cytidine deaminases. However, various species of lentiviral Vif proteins have evolved different strategies for overcoming host APOBEC3. Whether different species of lentiviral Vif proteins still preserve certain common features has not been reported. Here, we show for the first time that diverse lentiviral Vif molecules maintain the ability to interact with the HIV-1 Gag precursor (Pr55(Gag)) polyprotein. Surprisingly, BIV Vif, but not HIV-1 Vif, interfered with HIV-1 production and viral infectivity even in the absence of APOBEC3. Further analysis revealed that BIV Vif demonstrated an enhanced interaction with Pr55(Gag) when compared to HIV-1 Vif, and BIV Vif defective for the Pr55(Gag) interaction lost its ability to inhibit HIV-1. The C-terminal region of CA and the p2 region of Pr55(Gag), which are important for virus assembly and maturation, were involved in the interaction. Transduction of CD4(+) T cells with BIV Vif blocked HIV-1 replication. Thus, the conserved Vif-Pr55(Gag) interaction provides a potential target for the future development of anti-viral strategies.

Concepts: HIV, DNA, Protein, Gene, Molecular biology, Virology, Open reading frame, Viral infectivity factor


The human cytidine deaminase APOBEC3G (A3G) is a potent inhibitor of the HIV-1 virus in the absence of viral infectivity factor (Vif). The molecular mechanism of A3G antiviral activity is primarily attributed to deamination of single-stranded DNA (ssDNA); however the non-deamination mechanism also contributes to HIV-1 restriction. The interaction of A3G with ssDNA and RNA is reguired for its antiviral activity. Here we used atomic force microscopy (AFM) to directly visualize A3G-RNA and A3G-ssDNA complexes and compare to each other. Our results showed that A3G in A3G-RNA complexes exists primerely in monomeric-dimeric states, similar to its stoichiometry in complexes with ssDNA. New A3G-RNA complexes in which A3G binds to two RNA molecules were identified. These data suggest the existence of two separate RNA binding sites on A3G. Such complexes were not observed with ssDNA substrates. Time-lapse high-speed Atomic force microscopy (HS-AFM) was applied to characterize the dynamics of the complexes. The data revealed that the two RNA binding sites have different affinities to A3G. Based on the obtained results, a model for the interaction of A3G with RNA is proposed.

Concepts: DNA, Protein, Gene, Molecular biology, Virus, RNA, Cytosine, Viral infectivity factor


The essential HIV-1 viral infectivity factor (Vif) allows productive infection of non-permissive cells expressing cytidine deaminases APOBEC3G (A3G) and A3F by decreasing their cellular level, and preventing their incorporation into virions. Unlike the Vif-induced degradation of A3G, the functional role of the inhibition of A3G translation by Vif remained unclear. Here, we show that two stem-loop structures within the 5'-untranslated region of A3G mRNA are crucial for translation inhibition by Vif in cells, and most Vif alleles neutralize A3G translation efficiently. Interestingly, K26R mutation in Vif abolishes degradation of A3G by the proteasome but has no effect at the translational level, indicating these two pathways are independent. These two mechanisms, proteasomal degradation and translational inhibition, similarly contribute to decrease the cellular level of A3G by Vif and to prevent its incorporation into virions. Importantly, inhibition of A3G translation is sufficient to partially restore viral infectivity in the absence of proteosomal degradation. These findings demonstrate that HIV-1 has evolved redundant mechanisms to specifically inhibit the potent antiviral activity of A3G.

Concepts: DNA, Gene, Genetics, Gene expression, Evolution, RNA, Viral infectivity factor, APOBEC3G