Human Apolipoprotein B mRNA-editing Enzyme-catalytic Polypeptide-like 3G (APOBEC3G) Is Incorporated into HIV-1 Virions through Interactions with Viral and Nonviral RNAs

Abstract

Apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G (APOBEC3G) is a host cytidine deaminase that is packaged into virions and confers resistance to retroviral infection. APOBEC3G deaminates deoxycytidines in minus strand DNA to deoxyuridines, resulting in G to A hypermutation and viral inactivation. Human immunodeficiency virus type 1 (HIV-1) virion infectivity factor counteracts the antiviral activity of APOBEC3G by inducing its proteosomal degradation and preventing virion incorporation. To elucidate the mechanism of viral suppression by APOBEC3G, we developed a sensitive cytidine deamination assay and analyzed APOBEC3G virion incorporation in a series of HIV-1 deletion mutants. Virus-like particles derived from constructs in which pol, env, and most of gag were deleted still contained high levels of cytidine deaminase activity; in addition, coimmunoprecipitation of APOBEC3G and HIV-1 Gag in the presence and absence of RNase A indicated that the two proteins do not interact directly but form an RNase-sensitive complex. Viral particles lacking HIV-1 genomic RNA which were generated from the gag-pol expression constructs pCHelp and pSYNGP packaged APOBEC3G at 30 -40% of the wild-type level, indicating that interactions with viral RNA are not necessary for incorporation. In addition, viral particles produced from an nucleocapsid zinc finger mutant contained ϳ1% of the viral genomic RNA but ϳ30% of the cytidine deaminase activity. The reduction in APOBEC3G incorporation was equivalent to the reduction in the total RNA present in the nucleocapsid mutant virions. These results indicate that interactions with viral proteins or viral genomic RNA are not essential for APOBEC3G incorporation and suggest that APOBEC3G interactions with viral and nonviral RNAs that are packaged into viral particles are sufficient for APOBEC3G virion incorporation.

Enzymatically Active APOBEC3G Is Required for Efficient Inhibition of Human Immunodeficiency Virus Type 1

“…APO3G is packaged into viruses or virus-like particles in a process that involves nucleocapsid protein and viral genomic RNA (4,5,7,17,21,32,39,44). Moreover, we previously reported that APO3G packaged in the presence of viral genomic RNA stably associates with viral nucleoprotein complexes and is resistant to detergent treatment while APO3G packaged in the absence of viral genomic RNA is detergent sensitive and presumably not core associated (17).…”

Resistance of human T cell leukemia virus type 1 to APOBEC3G restriction is mediated by elements in nucleocapsid

“…It is relevant that the C terminus of HTLV-1 NC contributes to the exclusion of hA3G from VLPs, because it is NC that brings viral and cellular RNAs into the virion. It is currently believed that both NC and RNA are necessary for incorporation of hA3G into virions (19,20,22,(32)(33)(34), but it is unclear whether hA3G is simply tethered to RNA (21,22,34) or interacts with a NC-RNA complex (11). Deleting 20 aa near the C terminus of HTLV-1 NC (NCDC mutant) did not alter the amount of viral RNA in virions but did increase the amount of hA3G that was packaged.…”

“…Although cytosine deaminase activity appears to correlate with antiviral activity of hA3G, there is evidence that other factors may contribute to antiviral activity, and it is still unclear which stage of the virus infectious cycle is primarily affected (16)(17)(18). The exact mechanism by which hA3G is packaged into virions is also unresolved but it does appear to require both RNA and viral nucleocapsid (NC) protein (19)(20)(21)(22). HIV-1 counteracts the effects of hA3G with the accessory protein Vif, whose primary function is to target hA3G for proteasomal degradation, thereby preventing encapsidation of hA3G into virus particles (23)(24)(25)(26)(27)(28).…”

APOBEC proteins and intrinsic resistance to HIV-1 infection

“…The current consensus view is that A3G packaging is determined by specific interactions between the N-terminal CDA domain of A3G and the nucleocapsid (NC) region of Gag that are also dependent upon the binding of A3G to RNA (Luo et al 2004;Schafer et al 2004;Svarovskaia et al 2004;Zennou et al 2004;Khan et al 2005;Navarro et al 2005;Burnett & Spearman 2007;Bogerd & Cullen 2008). Accordingly, A3G is not encapsidated by either HIV-1 virus-like particles that lack NC domains (Schafer et al 2004;Zennou et al 2004) or other retroviruses whose Gag proteins do not form complexes with A3G (Doehle et al 2006).…”