Loss of tetherin antagonism by Nef impairs SIV replication during acute infection of rhesus macaques

Abstract

Most simian immunodeficiency viruses use Nef to counteract the tetherin proteins of their nonhuman primate hosts. Nef also downmodulates cell-surface CD4 and MHC class I (MHC I) molecules and enhances viral infectivity by counteracting SERINC5. We previously demonstrated that tetherin antagonism by SIV Nef is genetically separable from CD4- and MHC I-downmodulation. Here we show that disruption of tetherin antagonism by Nef impairs virus replication during acute SIV infection of rhesus macaques. A combination of mutations was introduced into the SIV(mac)239 genome resulting in three amino acid substitutions in Nef that impair tetherin antagonism, but not CD3-, CD4- or MHC I-downmodulation. Further characterization of this mutant (SIV(mac)239(AAA)) revealed that these changes also result in partial sensitivity to SERINC5. Separate groups of four rhesus macaques were infected with either wild-type SIV(mac)239 or SIV(mac)239(AAA), and viral RNA loads in plasma and sequence changes in the viral genome were monitored. Viral loads were significantly lower during acute infection in animals infected with SIV(mac)239(AAA) than in animals infected with wild-type SIV(mac)239. Sequence analysis of the virus population in plasma confirmed that the substitutions in Nef were retained during acute infection; however, changes were observed by week 24 post-infection that fully restored anti-tetherin activity and partially restored anti-SERINC5 activity. These observations reveal overlap in the residues of SIV Nef required for counteracting tetherin and SERINC5 and selective pressure to overcome these restriction factors in vivo. Author summary The SIV Nef protein enhances virus replication by removing certain cellular proteins from the surface of infected cells. Whereas the downmodulation of tetherin enhances virus release from infected cells, the downmodulation of SERINC5, and to a lesser extent SERINC3, increases viral infectivity. Although these Nef-mediated effects on the host cell have been demonstrated in vitro, the relevance of these activities to virus replication in primates has not been directly assessed. We therefore introduced mutations into SIV that render Nef ineffective against tetherin and SERINC5. Rhesus macaques infected with this virus exhibited lower acute viremia compared to animals infected with wild-type SIV. We also observed a partial recovery of these functions as a result of the accumulation of compensatory mutations in Nef. This study therefore demonstrates that overcoming restriction by tetherin and SERINC5 is important for efficient replication of immunodeficiency viruses in their infected hosts.