It has become clear in the last few years that human cells contain an enzyme, APOBEC3G (hA3G), that induces profound resistance to infection by certain retroviruses. hA3G protein possesses cytidine deaminase activity, and one mechanism responsible for its antiviral effects is deamination of cytosine residues in minus-strand DNA, producing G-to-A mutation in the coding strand of the provirus. HIV-1 encodes a protein, Vif, that blocks the effects of hA3G by binding to it and promoting its degradation in the proteasome. Despite several years of intensive study, the mechanism of hA3G incorporation into virions and the existence of antiviral effects other than deamination are still unresolved questions. _____While the interaction of human A3G with HIV-1 has been a central object of investigation in many laboratories, it is clear that other members of the APOBEC3 family can also have antiviral effects, that APOBEC3 family members are present in many mammalian species, and that different viruses have distinct patterns of sensitivity to the different APOBEC3 isoforms. Mice contain only one family member, APOBEC3 (mA3). it was originally reported that MLVs are resistant to mA3 because they do not incorporate it into assembling virions, whereas other studies indicate that mA3 is incorporated into MLV particles without a significant antiviral effect. _____In collaboration with the late Dr. David Derse, we re-examined the response of MLV and MLV-derived vectors to mA3, along with their sensitivity to hA3G. We found, contrary to the published reports, that MLV and related vectors are sensitive to mA3, although they are considerably more sensitive to hA3G. Other experiments showed that the potency of mA3 against delta-vif HIV-1 is equal to that of hA3G. We have been unable to detect G:A hypermutation induced by mA3 following MLV infections, although high levels of the mutations are observed with MLV inactivated by hA3G. This observation supports the concept that G:A hypermutation is not the only mechanism by which APOBEC proteins interfere with retroviral infections. In contrast, mA3 does induce G:A hypermutation in delta-vif HIV-1. _____Taken together, the data show that MLV is partially resistant to the antiviral activity of mA3. MLV is a simple retrovirus, encoding only the three polyproteins that are assembled to form infectious progeny virions. Thus, it would be of considerable interest to determine the mechanism of its resistance to mA3. We have produced enzymatically active mA3 in insect cells and have characterized its biochemical properties. We have also detected the deaminase activity of mA3 in MLV virions, despite the fact that mA3 is incapable of inducing deamination in the DNA products of these virions. _____Recent Accomplishments and Current Research: _____a. Analysis of mechanisms of MLV inactivation by, and resistance to, mA3. Moloney MLV is largely resistant to mouse APOBEC3 (mA3) and completely resistant to mA3-induced hypermutation. We replaced the gag gene of Moloney MLV with that of an endogenous polytropic MLV. The response of the resulting virus to mA3 is similar to that of Moloney MLV. Our data also show that mA3 blocks MLV infection before or at the beginning of reverse transcription. _____b. Biochemical studies on recombinant mA3. We are purifying recombinant, enzymatically active mA3 protein. We have characterized it biochemically and have looked for interactions with Moloney MLV proteins that might be involved in the mA3 resistance of Moloney MLV. We are continuing to study the effects of mA3 and its possible interactions with MLV glyco-Gag. _____[Corresponds to Rein Project 3 in the October 2011 site visit report of the HIV Drug Resistance Program (renamed HIV Dynamics and Replication Program in 2015)]
| Nair, Smita; Sanchez-Martinez, Silvia; Ji, Xinhua et al. (2014) Biochemical and biological studies of mouse APOBEC3. J Virol 88:3850-60 |
| Nair, Smita; Rein, Alan (2014) Antiretroviral restriction factors in mice. Virus Res 193:130-4 |
| Stavrou, Spyridon; Nitta, Takayuki; Kotla, Swathi et al. (2013) Murine leukemia virus glycosylated Gag blocks apolipoprotein B editing complex 3 and cytosolic sensor access to the reverse transcription complex. Proc Natl Acad Sci U S A 110:9078-83 |
| Sanchez-Martinez, Silvia; Aloia, Amanda L; Harvin, Demetria et al. (2012) Studies on the restriction of murine leukemia viruses by mouse APOBEC3. PLoS One 7:e38190 |
| Rein, Alan; Datta, Siddhartha A K; Jones, Christopher P et al. (2011) Diverse interactions of retroviral Gag proteins with RNAs. Trends Biochem Sci 36:373-80 |
