Newly synthesized APOBEC3G is incorporated into HIV virions, inhibited by HIV RNA, and subsequently activated by RNase H

Department of Medicine, University of California, San Francisco, San Francisco, California, United States
PLoS Pathogens (Impact Factor: 7.56). 03/2007; 3(2):e15. DOI: 10.1371/journal.ppat.0030015
Source: PubMed


Author Summary

APOBEC3G (A3G) is a cellular enzyme that promotes DNA mutagenesis and can restrict infection by HIV-1. However, HIV counters the antiviral effects of A3G through the action of its Vif protein. In the absence of Vif, A3G is effectively incorporated into virions, where it mutagenizes the first DNA copy (cDNA) generated during reverse transcription of the viral RNA genome. A3G also appears to be able to inhibit HIV via nonenzymatic mechanisms. A3G and related deoxycytidine deaminases can also inhibit the growth of retroviruses other than HIV and protect the cellular genome from endogenous mobile retroelements. In this study, we analyzed the recruitment and enzymatic activity of A3G incorporated into HIVΔVif virions. Unexpectedly, we found that the binding of A3G to viral genomic RNA led to inactivation of the enzyme. However, latent A3G was ultimately activated through the action of HIV RNase H, which degrades the RNA genome during reverse transcription. These findings highlight an unexpected interplay between a host enzyme and HIV, where the antiviral enzymatic activity of the host factor (A3G) is dependent on the action of an essential HIV enzyme (RNase H). The strong interaction with viral RNA also suggests a potential mechanism by which A3G could exert antiviral activity in the absence of enzymatic activity, by physically impeding reverse transcription.

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    • "Vif was not associated with these high molecular mass A3G forms, but was required for their formation (Goila-Gaur et al., 2008). Although A3G regularly forms high molecular mass complexes in cells, which are less likely to be packaged into virions, Vif can induce an even higher molecular weight form of A3G (Soros et al., 2007; Goila-Gaur et al., 2008). Moreover, studies with an A3G C97A mutant that is resistant to Vif-mediated degradation suggested that Vif-mediated degradation and inhibition of packaging are two distinct properties of A3G since the A3G C97A mutant was encapsidated less well in the presence of Vif (Opi et al., 2007). "
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    ABSTRACT: The APOBEC3 restriction factors are a family of deoxycytidine deaminases that are able to suppress replication of viruses with a single-stranded DNA intermediate by inducing mutagenesis and functional inactivation of the virus. Of the seven human APOBEC3 enzymes, only APOBEC3-D, -F, -G, and -H appear relevant to restriction of HIV-1 in CD4+ T cells and will be the focus of this review. The restriction of HIV-1 occurs most potently in the absence of HIV-1 Vif that induces polyubiquitination and degradation of APOBEC3 enzymes through the proteasome pathway. To restrict HIV-1, APOBEC3 enzymes must be encapsidated into budding virions. Upon infection of the target cell during reverse transcription of the HIV-1 RNA into (-)DNA, APOBEC3 enzymes deaminate cytosines to form uracils in single-stranded (-)DNA regions. Upon replication of the (-)DNA to (+)DNA, the HIV-1 reverse transcriptase incorporates adenines opposite to the uracils thereby inducing C/G to T/A mutations that can functionally inactivate HIV-1. APOBEC3G is the most studied APOBEC3 enzyme and it is known that Vif attempts to thwart APOBEC3 function not only by inducing its proteasomal degradation but also by several degradation-independent mechanisms, such as inhibiting APOBEC3G virion encapsidation, mRNA translation, and for those APOBEC3G molecules that still become virion encapsidated, Vif can inhibit APOBEC3G mutagenic activity. Although most Vif variants can induce efficient degradation of APOBEC3-D, -F, and -G, there appears to be differential sensitivity to Vif-mediated degradation for APOBEC3H. This review examines APOBEC3-mediated HIV restriction mechanisms, how Vif acts as a substrate receptor for a Cullin5 ubiquitin ligase complex to induce degradation of APOBEC3s, and the determinants and functional consequences of the APOBEC3 and Vif interaction from a biological and biochemical perspective.
    Full-text · Article · Aug 2014 · Frontiers in Microbiology
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    • "In agreement with this hypothesis, hA3G Y124A that is deficient in its ability to form the RA LMM and HMM complexes (the precursor and mature forms), is unable to be packaged into virions, even though it is able to interacts with Gag as efficiently as wild type hA3G (Figure 5). Furthermore, the moving of newly-synthesized hA3G to the lipid raft domains to rapidly form the RA LMM is consistent with a previous finding that hA3G is incorporated into virion shortly after its synthesis in cytoplasm [17]. These data together indicate that the RA LMM hA3G complex acts as the cellular source for its virion encapsidation. "
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    ABSTRACT: The incorporation of human APOBEC3G (hA3G) into HIV is required for exerting its antiviral activity, therefore the mechanism underlying hA3G virion encapsidation has been investigated extensively. hA3G was shown to form low-molecular-mass (LMM) and high-molecular-mass (HMM) complexes. The function of different forms of hA3G in its viral incorporation remains unclear. In this study, we investigated the subcellular distribution and lipid raft association of hA3G using subcellular fractionation, membrane floatation assay and pulse-chase radiolabeling experiments respectively, and studied the correlation between the ability of hA3G to form the different complex and its viral incorporation. Our work herein provides evidence that the majority of newly-synthesized hA3G interacts with membrane lipid raft domains to form Lipid raft-associated hA3G (RA hA3G), which serve as the precursor of mature HMM hA3G complex, while a minority of newly-synthesized hA3G remains in the cytoplasm as a soluble LMM form. The distribution of hA3G among the soluble LMM form, the RA LMM form and the mature forms of HMM is regulated by a mechanism involving the N-terminal part of the linker region and the C-terminus of hA3G. Mutagenesis studies reveal a direct correlation between the ability of hA3G to form the RA LMM complex and its viral incorporation. Together these data suggest that the Lipid raft-associated LMM A3G complex functions as the cellular source of viral hA3G.
    Full-text · Article · Oct 2013 · PLoS ONE
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    • "Noteworthy, the knockdown of A3G in unstimulated CD4+ T cells does not turn them permissive to infection, suggesting that the presence of LMM A3G in the cells is not the unique determinant for their resistance to HIV-1 [62, 63]. Moreover, LMM A3G is preferentially packaged into HIV-1 particles [56, 64–66]. Finally, HMM A3G is also able to interact with and sequester Alu RNA elements, inhibiting their transposition and evidencing the role of different A3G molecular forms in the restriction of retroelements [67]. "
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    ABSTRACT: The APOBEC family of proteins comprises deaminase enzymes that edit DNA and/or RNA sequences. The APOBEC3 subgroup plays an important role on the innate immune system, acting on host defense against exogenous viruses and endogenous retroelements. The role of APOBEC3 proteins in the inhibition of viral infection was firstly described for HIV-1. However, in the past few years many studies have also shown evidence of APOBEC3 action on other viruses associated with human diseases, including HTLV, HCV, HBV, HPV, HSV-1, and EBV. APOBEC3 inhibits these viruses through a series of editing-dependent and independent mechanisms. Many viruses have evolved mechanisms to counteract APOBEC effects, and strategies that enhance APOBEC3 activity constitute a new approach for antiviral drug development. On the other hand, novel evidence that editing by APOBEC3 constitutes a source for viral genetic diversification and evolution has emerged. Furthermore, a possible role in cancer development has been shown for these host enzymes. Therefore, understanding the role of deaminases on the immune response against infectious agents, as well as their role in human disease, has become pivotal. This review summarizes the state-of-the-art knowledge of the impact of APOBEC enzymes on human viruses of distinct families and harboring disparate replication strategies.
    Full-text · Article · Jun 2013
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