Lily I. Wu

HIV-1 Infection Requires a Functional Integrase NLS

HIV-1 is able to infect nondividing cells productively in part because the postentry viral nucleoprotein complexes are actively imported into the nucleus. In this manuscript, we identify a novel nuclear localization signal (NLS) in the viral integrase (IN) protein that is essential for virus replication in both dividing and nondividing cells. The IN NLS stimulates the efficient nuclear accumulation of viral DNA as well as virion-derived IN protein during the initial stages of infection but is dispensable for catalytic function. Because this NLS is required for infection irrespective of target cell proliferation, we suggest that interactions between uncoated viral nucleoprotein complexes and the host cell nuclear import machinery are critical for HIV-1 infection of all cells.

High-Frequency Persistence of an Impaired Allele of the Retroviral Defense Gene TRIM5α in Humans

The intracellular TRIM5alpha protein successfully inhibits HIV-1 infection in rhesus monkeys, but not in humans . A few amino acids in the virus-interacting SPRY domain were found to be responsible for most of this anti-viral specificity , raising the possibility that genetic variation among humans could result in TRIM5alpha proteins with a spectrum of potencies. We found several nonsynonymous SNPs at the human TRIM5 locus, but only one of these (H43Y) was found to have a significant functional consequence. We demonstrate that H43Y impairs TRIM5alpha restriction of two distantly related retroviruses. H43Y lies in the RING domain of TRIM5alpha and may negatively affect its putative E3 ubiquitin ligase activity. This detrimental allele dates back to before the African diaspora and is found at a frequency of 43% in indigenous Central and South Americans. We suggest that relaxed constraint due to a recent period of low retroviral challenge has allowed the deleterious H43Y mutation to persist and even to expand after the bottleneck that occurred upon human migration to the New World. The unexpectedly high frequency of an impaired retroviral restriction allele among humans is likely to have a significant impact on our ability to ward off future retroviral challenges.

The Range of Human APOBEC3H Sensitivity to Lentiviral Vif Proteins

ABSTRACT The APOBEC3H gene is polymorphic in humans, with four major population-dependent haplotypes that encode proteins with different levels of antiviral activity. Haplotype II, present most frequently in African populations, encodes the most stable protein and is most active against human immunodeficiency virus type 1 (HIV-1). In contrast to human APOBEC3G, which can be completely counteracted by HIV-1 Vif, the protein encoded by APOBEC3H haplotype II is only partially sensitive to Vif, while the protein encoded by APOBEC3H haplotype I is completely resistant to HIV-1 Vif. We mapped a residue on APOBEC3H that determines this partial Vif sensitivity. However, it is unclear how HIV-1 can replicate in vivo without the ability to neutralize APOBEC3H antiviral activity. In order to directly address this question, we cloned vif genes from HIV-1-infected individuals with different APOBEC3H genotypes and tested them for their ability to inhibit human APOBEC3H. We found that while the APOBEC3H genotype of infected individuals significantly influences the activity of Vif encoded by their virus, none of the Vif variants tested can completely neutralize APOBEC3H as well as they neutralize APOBEC3G. Consistent with this genetic result, APOBEC3H protein expression in human peripheral blood mononuclear cells was below our limit of detection using newly developed antibodies against the endogenous protein. These results demonstrate that human APOBEC3H is not as strong of a selective force for current HIV-1 infections as human APOBEC3G.

Birth, decay, and reconstruction of an ancient TRIMCyp gene fusion in primate genomes

TRIM5 is a host antiviral gene with an evolutionary history of genetic conflict with retroviruses. The TRIMCyp gene encodes a protein fusion of TRIM5 effector domains with the capsid-binding ability of a retrotransposed CyclophilinA (CypA), resulting in novel antiviral specificity against lentiviruses. Previous studies have identified two independent primate TRIMCyp fusions that evolved within the past 6 My. Here, we describe an ancient primate TRIMCyp gene (that we call TRIMCypA3), which evolved in the common ancestor of simian primates 43 Mya. Gene reconstruction shows that CypA3 encoded an intact, likely active, TRIMCyp antiviral gene, which was subject to selective constraints for at least 10 My, followed by pseudogenization or loss in all extant primates. Despite its decayed status, we found TRIMCypA3 gene fusion transcripts in several primates. We found that the reconstructed “newly born” TrimCypA3 encoded robust and broad retroviral restriction activity but that this broad activity was lost via eight amino acid changes over the course of the next 10 My. We propose that TRIMCypA3 arose in response to a viral pathogen encountered by ancestral primates but was subsequently pseudogenized or lost due to a lack of selective pressure. Much like imprints of ancient viruses, fossils of decayed genes, such as TRIMCypA3, provide unique and specific insight into paleoviral infections that plagued primates deep in their evolutionary history.

The function and evolution of the restriction factor viperin in primates was not driven by lentiviruses

BackgroundViperin, also known as RSAD2, is an interferon-inducible protein that potently restricts a broad range of different viruses such as influenza, hepatitis C virus, human cytomegalovirus and West Nile virus. Viperin is thought to affect virus budding by modification of the lipid environment within the cell. Since HIV-1 and other retroviruses depend on lipid domains of the host cell for budding and infectivity, we investigated the possibility that Viperin also restricts human immunodeficiency virus and other retroviruses.ResultsLike other host restriction factors that have a broad antiviral range, we find that viperin has also been evolving under positive selection in primates. The pattern of positive selection is indicative of Viperins escape from multiple viral antagonists over the course of primate evolution. Furthermore, we find that Viperin is interferon-induced in HIV primary target cells. We show that exogenous expression of Viperin restricts the LAI strain of HIV-1 at the stage of virus release from the cell. Nonetheless, the effect of Viperin restriction is highly strain-specific and does not affect most HIV-1 strains or other retroviruses tested. Moreover, knockdown of endogenous Viperin in a lymphocytic cell line did not significantly affect the spreading infection of HIV-1.ConclusionDespite positive selection having acted on Viperin throughout primate evolution, our findings indicate that Viperin is not a major restriction factor against HIV-1 and other retroviruses. Therefore, other viral lineages are likely responsible for the evolutionary signatures of positive selection in viperin among primates.

The Role of the Antiviral APOBEC3 Gene Family in Protecting Chimpanzees against Lentiviruses from Monkeys

Cross-species transmissions of viruses from animals to humans are at the origin of major human pathogenic viruses. While the role of ecological and epidemiological factors in the emergence of new pathogens is well documented, the importance of host factors is often unknown. Chimpanzees are the closest relatives of humans and the animal reservoir at the origin of the human AIDS pandemic. However, despite being regularly exposed to monkey lentiviruses through hunting, chimpanzees are naturally infected by only a single simian immunodeficiency virus, SIVcpz. Here, we asked why chimpanzees appear to be protected against the successful emergence of other SIVs. In particular, we investigated the role of the chimpanzee APOBEC3 genes in providing a barrier to infection by most monkey lentiviruses. We found that most SIV Vifs, including Vif from SIVwrc infecting western-red colobus, the chimpanzee’s main monkey prey in West Africa, could not antagonize chimpanzee APOBEC3G. Moreover, chimpanzee APOBEC3D, as well as APOBEC3F and APOBEC3H, provided additional protection against SIV Vif antagonism. Consequently, lentiviral replication in primary chimpanzee CD4+ T cells was dependent on the presence of a lentiviral vif gene that could antagonize chimpanzee APOBEC3s. Finally, by identifying and functionally characterizing several APOBEC3 gene polymorphisms in both common chimpanzees and bonobos, we found that these ape populations encode APOBEC3 proteins that are uniformly resistant to antagonism by monkey lentiviruses.

An Evolutionary Screen Highlights Canonical and Noncanonical Candidate Antiviral Genes within the Primate TRIM Gene Family

Recurrent viral pressure has acted on host-encoded antiviral genes during primate and mammalian evolution. This selective pressure has resulted in dramatic episodes of adaptation in host antiviral genes, often detected via positive selection. These evolutionary signatures of adaptation have the potential to highlight previously unrecognized antiviral genes (also called restriction factors). Although the TRIM multigene family is recognized for encoding several bona fide restriction factors (e.g., TRIM5alpha), most members of this expansive gene family remain uncharacterized. Here, we investigated the TRIM multigene family for signatures of positive selection to identify novel candidate antiviral genes. Our analysis reveals previously undocumented signatures of positive selection in 17 TRIM genes, 10 of which represent novel candidate restriction factors. These include the unusual TRIM52 gene, which has evolved under strong positive selection despite its encoded protein lacking a putative viral recognition (B30.2) domain. We show that TRIM52 arose via gene duplication from the TRIM41 gene. Both TRIM52 and TRIM41 have dramatically expanded RING domains compared with the rest of the TRIM multigene family, yet this domain has evolved under positive selection only in primate TRIM52, suggesting that it represents a novel host–virus interaction interface. Our evolutionary-based screen not only documents positive selection in known TRIM restriction factors but also highlights candidate novel restriction factors, providing insight into the interfaces of host–pathogen interactions mediated by the TRIM multigene family.

A Single Nucleotide Polymorphism in Human APOBEC3C Enhances Restriction of Lentiviruses

Humans express seven human APOBEC3 proteins, which can inhibit viruses and endogenous retroelements through cytidine deaminase activity. The seven paralogs differ in the potency of their antiviral effects, as well as in their antiviral targets. One APOBEC3, APOBEC3C, is exceptional as it has been found to only weakly block viruses and endogenous retroelements compared to other APOBEC3s. However, our positive selection analyses suggest that APOBEC3C has played a role in pathogen defense during primate evolution. Here, we describe a single nucleotide polymorphism in human APOBEC3C, a change from serine to isoleucine at position 188 (I188) that confers potent antiviral activity against HIV-1. The gain-of-function APOBEC3C SNP results in increased enzymatic activity and hypermutation of target sequences when tested in vitro, and correlates with increased dimerization of the protein. The I188 is widely distributed in human African populations, and is the ancestral primate allele, but is not found in chimpanzees or gorillas. Thus, while other hominids have lost activity of this antiviral gene, it has been maintained, or re-acquired, as a more active antiviral gene in a subset of humans. Taken together, our results suggest that APOBEC3C is in fact involved in protecting hosts from lentiviruses.