Date Published: July 1, 2019
Publisher: Public Library of Science
Author(s): Amit Sharma, Richard N. McLaughlin, Ryan S. Basom, Caroline Kikawa, Molly OhAinle, Jacob S. Yount, Michael Emerman, Julie Overbaugh, David T. Evans.
HIV-1 does not persistently infect macaques due in part to restriction by several macaque host factors. This has been partially circumvented by generating chimeric SIV/HIV-1 viruses (SHIVs) that encode SIV antagonist of known restriction factors. However, most SHIVs replicate poorly in macaques unless they are further adapted in culture and/or macaques (adapted SHIVs). Therefore, development of SHIVs encoding HIV-1 sequences derived directly from infected humans without adaptation (unadapted SHIVs) has been challenging. In contrast to the adapted SHIVs, the unadapted SHIVs have lower replication kinetics in macaque lymphocytes and are sensitive to type-1 interferon (IFN). The HIV-1 Envelope (Env) in the chimeric virus determines both the reduced replication and the IFN-sensitivity differences. There is limited information on macaque restriction factors that specifically limit replication of the more biologically relevant, unadapted SHIV variants. In order to identify the IFN-induced host factor(s) that could contribute to the inhibition of SHIVs in macaque lymphocytes, we measured IFN-induced gene expression in immortalized pig-tailed macaque (Ptm) lymphocytes using RNA-Seq. We found 147 genes that were significantly upregulated upon IFN treatment in Ptm lymphocytes and 31/147 were identified as genes that encode transmembrane helices and thus are likely present in membranes where interaction with viral Env is plausible. Within this group of upregulated genes with putative membrane-localized proteins, we identified several interferon-induced transmembrane protein (IFITM) genes, including several previously uncharacterized Ptm IFITM3-related genes. An evolutionary genomic analysis of these genes suggests the genes are IFITM3 duplications not found in humans that are both within the IFITM locus and also dispersed elsewhere in the Ptm genome. We observed that Ptm IFITMs are generally packaged at higher levels in unadapted SHIVs when compared to adapted SHIVs. CRISPR/Cas9-mediated knockout of Ptm IFITMs showed that depletion of IFITMs partially rescues the IFN sensitivity of unadapted SHIV. Moreover, we found that the depletion of IFITMs also increased replication of unadapted SHIV in the absence of IFN treatment, suggesting that Ptm IFITMs are likely important host factors that limit replication of unadapted SHIVs. In conclusion, this study shows that Ptm IFITMs selectively restrict replication of unadapted SHIVs. These findings suggest that restriction factors including IFITMs vary in their potency against different SHIV variants and may play a role in selecting for viruses that adapt to species-specific restriction factors.
The macaque models of HIV-1 infection have been critical to the understanding of retroviral pathogenesis as well as for testing antiretroviral therapies and candidate vaccines for HIV-1. However, multiple species-specific host factors restrict HIV-1 replication in macaque cells . To overcome these restrictions chimeric SIV/HIV-1 viruses (SHIVs), which encode the SIV antagonists of known restriction factors are used to infect macaques to model HIV-1 infection. Existing SHIV/macaque models typically employ SHIVs that encode HIV-1 sequences from viruses that were adapted by viral passage in cell culture, and often these viruses are from chronic stages of infection. However, there is evidence that the chronic stage HIV-1 variants are distinct from HIV-1 variants that are selected for transmission in humans [2, 3]. In addition, SHIVs encoding HIV-1 sequences derived directly from humans typically require further adaptation in vitro in macaque cells and/or in vivo by serial macaque-passage  in order to obtain pathogenic viruses that establish persistent infection in macaques. These variant chimeric viruses used in the SHIV/macaque models are thus “adapted” SHIVs. We have previously determined that most circulating, transmitted HIV-1 Envelope (Env) variants, including the transmitted/founder variants, do not use macaque CD4 entry receptor efficiently  and thus SHIVs generated using these Envs replicate poorly in macaque cells. The adaptation of SHIV Env sequences in macaques increases replication and pathogenicity of SHIVs [5–11] but also leads to antigenic changes in Env that can limit their utility for vaccine and therapeutic approaches . SHIVs encoding circulating HIV-1 variants obtained directly from the newly infected patients without adaptation (termed unadapted SHIVs) that maintain the antigenic properties of the transmitted variants are desired as challenge viruses for vaccine and therapeutic studies. However, most attempts at generating these SHIVs have failed as unadapted SHIVs replicate poorly, if at all, in macaque cells and do not establish persistent infection . The virus-host dynamics that contribute to the differences in replication of adapted and unadapted SHIVs in macaques are not well defined.
Here we identify a novel example of cross-species restriction in which macaque-specific restriction factors selectively restrict replication of SHIV based on circulating, transmitted HIV-1 Env variant. In this study, we employed transcriptional profiling to define the repertoire of ISGs induced by IFNα in Ptm CD4+ lymphocytes. On the basis of this profiling, IFITMs, which have been implicated as anti-viral factors for a number of different viruses, including lentiviruses, were identified as a candidate restriction factors in these cells. We documented IFITM gene duplications in the macaque genomes and evaluated the ability of previously uncharacterized Ptm IFITMs to restrict SHIVs. We found that a prototypical, macaque-adapted SHIV is resistant to IFITM-mediated restriction, whereas a prototypical, unadapted SHIV, which encodes a circulating HIV-1 Env variant, is inhibited by Ptm IFITMs. Further, we demonstrate that IFITM virion incorporation tracks with the IFNα sensitivity of the SHIVs. For example, IFITMs are packaged at higher levels in IFNα-sensitive SHIVs when compared to IFNα-resistant SHIVs. Overall, our results suggest that the increased replication that results from adaptation of SHIVs in macaques may in part reflect an adaptation to an IFITM-mediated restriction.