Overcoming macaque Tetherin to generate an improved animal model for HIV/AIDS
Final Report Abstract
30 years after the discovery of HIV-1 as the causative agent of AIDS, the virus is still a major health problem around the world. There is no functional cure for the disease and an effective vaccine has not be developed to date. The development of a non-human primate model for HIV-1 infection is hampered by the presence of restriction factors that inhibit various steps of the viral life cycle and are part of the innate immune system. Pigtailed macaques distinguish themselves from other non-human primates, as they do not express a TRIM5α protein that is active against HIV-1. Thus, by exchanging the HIV-1 Vif protein with its counterpart from SIVMAC it has been possible to develop a simian tropic HIV-1 strain (stHIV) that overcomes macaque APOBEC3-mediated restriction and is able to replicate efficiently in pigtailed macaque PBMCs. Inoculation of animals with stHIV leads to high viral titers during acute infection, but replication is eventually controlled and the animals do not progress to disease. When comparing the replication of SIVMAC with stHIV in vitro we noticed that while SIVMAC is only marginally inhibited in its ability to replicate in the presence of IFN-α, stHIV replication was completely abrogated. These findings suggested that there are additional restriction factors upregulated by IFN-α that inhibit stHIV replication in macaques. Tetherin is a well-studied restriction factor that is highly induced by IFN-α and blocks the release of mature virus particles from the cell surface in a species-specific manner. HIV-1 uses its Vpu protein to counteract human Tetherin. However, HIV-1 Vpu is not able to counteract macaque Tetherin. Therefore, we hypothesized that overcoming macaque Tetherin would improve our current HIV-1 based animal model and would enable stHIV to replicate in pgt PBMC upon the induction of the antiviral state through IFN-α. We generated two novel variants of stHIV by exchanging the HIV-1 Vpu protein with the Vpu proteins from SIVDEN or SIVGSN. We demonstrated that when introduced into stHIV these Vpu proteins induce CD4 downregulation and are able to efficiently counteract macaque Tetherin. Furthermore, these viruses replicate in pigtailed macaque PBMC with an efficiency comparable to that of the original stHIV and SIVMAC. Surprisingly however, these constructs are not able to restore or improve replication of stHIV in pgt PBMC in the presence of IFN, indicating that additional factors are involved in the inhibition of stHIV upon IFN treatment. In contrast to HIV-1, SIVMAC expresses a Vpx protein, which enables the virus to overcome a block by SAMHD1 in primary macrophages or resting CD4+ T-cells. We were able to generate a novel stHIV variant that expresses and efficiently packages Vpx into mature virus particles. Although this new construct was able to efficiently replicate in pgt PBMC, the addition of IFN-α completely abgrogated its replication. By single-cycle infection experiments we could show that at least part of this unknown speciesspecific, IFN-α induced restriction of stHIV replication occurs early in the retroviral life cycle and by using chimeric viruses we could effectively exclude TRIM5 proteins and any other CA- targeting factors as mediators of the target cell species-dependent inhibitory activity of IFN-α. Thus we could show that overcoming known species-specific restriction factors in macaques was not sufficient for efficient replication of stHIV in pgt PBMC in the presence of IFN and demonstrated the existence of further undiscovered, IFN-α inducible antiretroviral factors.
Publications
- Adaptation to the Interferon-induced antiviral state by human and simian immunodeficiency viruses. J Virol. 2013 Mar;87(6):3549-60
Bitzegeio J, Sampias M, Bieniasz PD, Hatziioannou T
