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Human immunodeficiency virus type 1 (HIV-1), the causative agent of AIDS, originated from simian immunodeficiency virus from chimpanzees (SIVcpz), the precursor of the human virus, approximately 100 years ago. This indicates that HIV-1 has emerged through the cross-species transmission of SIVcpz from chimpanzees to humans. However, it remains unclear how SIVcpz has evolved into pandemic HIV-1 in humans. To address this question, we inoculated three SIVcpz strains (MB897, EK505, and MT145), four pandemic HIV-1 strains (NL4-3, NLCSFV3, JRCSF, and AD8), and two nonpandemic HIV-1 strains (YBF30 and DJO0131). Humanized mice infected with SIVcpz strain MB897, a virus phylogenetically similar to pandemic HIV-1, exhibited a peak viral load comparable to that of mice infected with pandemic HIV-1, while peak viral loads of mice infected with SIVcpz strain EK505 or MT145 as well as nonpandemic HIV-1 strains were significantly lower. These results suggest that SIVcpz strain MB897 is preadapted to humans, unlike the other SIVcpz strains. Moreover, viral RNA sequencing of MB897-infected humanized mice identified a nonsynonymous mutation in env , a G413R substitution in gp120. The infectivity of the gp120 G413R mutant of MB897 was significantly higher than that of parental MB897. Furthermore, we demonstrated that the gp120 G413R mutant of MB897 augments the capacity for viral replication in both in vitro cell cultures and humanized mice. Taken together, this is the first experimental investigation to use an animal model to demonstrate a gain-of-function evolution of SIVcpz into pandemic HIV-1. IMPORTANCE From the mid-20th century, humans have been exposed to the menace of infectious viral diseases, such as severe acute respiratory syndrome coronavirus, Ebola virus, and Zika virus. These outbreaks of emerging/reemerging viruses can be triggered by cross-species viral transmission from wild animals to humans, or zoonoses. HIV-1, the causative agent of AIDS, emerged by the cross-species transmission of SIVcpz, the HIV-1 precursor in chimpanzees, around 100 years ago. However, the process by which SIVcpz evolved to become HIV-1 in humans remains unclear. Here, by using a hematopoietic stem cell-transplanted humanized-mouse model, we experimentally recapitulate the evolutionary process of SIVcpz to become HIV-1. We provide evidence suggesting that a strain of SIVcpz, MB897, preadapted to infect humans over other SIVcpz strains. We further demonstrate a gain-of-function evolution of SIVcpz in infected humanized mice. Our study reveals that pandemic HIV-1 has emerged through at least two steps: preadaptation and subsequent gain-of-function mutations., (Copyright © 2018 American Society for Microbiology.)

APOBEC3 (A3) family proteins are DNA cytosine deaminases recognized for contributing to HIV-1 restriction and mutation. Prior studies have demonstrated that A3D, A3F, and A3G enzymes elicit a robust anti-HIV-1 effect in cell cultures and in humanized mouse models. Human A3H is polymorphic and can be categorized into three phenotypes: stable, intermediate, and unstable. However, the anti-viral effect of endogenous A3H in vivo has yet to be examined. Here we utilize a hematopoietic stem cell-transplanted humanized mouse model and demonstrate that stable A3H robustly affects HIV-1 fitness in vivo. In contrast, the selection pressure mediated by intermediate A3H is relaxed. Intriguingly, viral genomic RNA sequencing reveled that HIV-1 frequently adapts to better counteract stable A3H during replication in humanized mice. Molecular phylogenetic analyses and mathematical modeling suggest that stable A3H may be a critical factor in human-to-human viral transmission. Taken together, this study provides evidence that stable variants of A3H impose selective pressure on HIV-1.

Background: Human endogenous retroviruses (HERVs), the remnants of ancient retroviral infections, constitute approximately 8% of human genomic DNA. Since HERV-K Gag expression is induced by HIV-1 Tat in T cells, induced HERV-K proteins could affect HIV-1 replication. Indeed, previously we showed that HERV-K Gag and HIV-1 Gag coassemble and that this appears to correlate with the effect of HERV-K Gag expression on HIV-1 particle release and its infectivity. We further showed that coassembly requires both MA and NC domains, which presumably serve as scaffolding for Gag via their abilities to bind membrane and RNA, respectively. Notably, however, despite possessing these abilities, MLV Gag failed to coassemble with HIV-1 Gag and did not affect assembly and infectivity of HIV-1 particles. It is unclear how the specificity of coassembly is determined., Results: Here, we showed that coexpression of HERV-K Gag with HIV-1 Gag changed size and morphology of progeny HIV-1 particles and severely diminished infectivity of such progeny viruses. We further compared HERV-K-MLV chimeric constructs to identify molecular determinants for coassembly specificity and for inhibition of HIV-1 release efficiency and infectivity. We found that the CA N-terminal domain (NTD) of HERV-K Gag is important for the reduction of the HIV-1 release efficiency, whereas both CA-NTD and major homology region of HERV-K Gag contribute to colocalization with HIV-1 Gag. Interestingly, these regions of HERV-K Gag were not required for reduction of progeny HIV-1 infectivity., Conclusions: Our results showed that HERV-K Gag CA is important for reduction of HIV-1 release and infectivity but the different regions within CA are involved in the effects on the HIV-1 release and infectivity. Altogether, these findings revealed that HERV-K Gag interferes the HIV-1 replication by two distinct molecular mechanisms.

Human immunodeficiency virus type 1 (HIV-1), the causative agent of acquired immunodeficiency syndrome, encodes four accessory genes, one of which is viral protein U (Vpu). Recently, the study of Vpu has been of great interest. For instance, various cellular proteins are degraded (e.g. CD4) and down-modulated (e.g. tetherin) by Vpu. Vpu also antagonizes the function of tetherin and inhibits NF-κB. Moreover, Vpu is a viroporin forming ion channels and may represent a promising target for anti-HIV-1 drugs. In this review, we summarize the domains/residues that are responsible for Vpu's functions, describe the current understanding of the role of Vpu in HIV-1-infected cells, and review the effect of Vpu on HIV-1 in replication and pathogenesis. Future investigations that simultaneously assess a combination of Vpu functions are required to clearly delineate the most important functions for viral replication. Impact statement Viral protein U (Vpu) is a unique protein encoded by human immunodeficiency virus type 1 (HIV-1) and related lentiviruses, playing multiple roles in viral replication and pathogenesis. In this review, we briefly summarize the most up-to-date knowledge of HIV-1 Vpu.

APOBEC3 (apolipoprotein B mRNA editing enzyme catalytic polypeptide-like 3) proteins are cellular DNA deaminases that restrict a broad spectrum of lentiviruses. This process is counteracted by Vif (viral infectivity factor) of lentiviruses, which binds APOBEC3s and promotes their degradation. CBF-β (core binding factor subunit β) is an essential co-factor for the function of human immunodeficiency virus type 1 Vif to degrade human APOBEC3s. However, the requirement for CBF-β in Vif-mediated degradation of other mammalian APOBEC3 proteins is less clear. Here, we determined the sequence of feline CBFB and performed phylogenetic analyses. These analyses revealed that mammalian CBFB is under purifying selection. Moreover, we demonstrated that CBF-β is dispensable for feline immunodeficiency virus Vif-mediated degradation of APOBEC3s of its host. These findings suggested that primate lentiviruses have adapted to use CBF-β, an evolutionary stable protein, to counteract APOBEC3 proteins of their hosts after diverging from other lentiviruses., (© 2015 The Authors.)

Human immunodeficiency virus type 1 (HIV-1) encodes four accessory genes: vif, vpu, vpr, and nef. Recent investigations using in vitro cell culture systems have shed light on the roles of these HIV-1 accessory proteins, Vif, Vpr, Vpu, and Nef, in counteracting, modulating, and evading various cellular factors that are responsible for anti-HIV-1 intrinsic immunity. However, since humans are the exclusive target for HIV-1 infection, conventional animal models are incapable of mimicking the dynamics of HIV-1 infection in vivo. Moreover, the effects of HIV-1 accessory proteins on viral infection in vivo remain unclear. To elucidate the roles of HIV-1 accessory proteins in the dynamics of viral infection in vivo, humanized mouse models, in which the mice are xenotransplanted with human hematopoietic stem cells, has been utilized. This review describes the current knowledge of the roles of HIV-1 accessory proteins in viral infection, replication, and pathogenicity in vivo, which are revealed by the studies using humanized mouse models.

Bimolecular fluorescence complementation (BiFC) and western blot analysis demonstrated that CCR5 exists as constitutive homo-oligomers, which was further enhanced by its antagonists such as maraviroc (MVC) and TAK-779. Staining by monoclonal antibodies recognizing different epitopes of CCR5 revealed that CCR5 oligomer was structurally different from the monomer. To determine which forms of CCR5 are well recognized by CCR5-using HIV-1 for the entry, BiFC-positive and -negative cell fractions in CD4-positive 293T cells were collected by fluorescent-activated cell sorter, and infected with luciferase-reporter HIV-1 pseudotyped with CCR5-using Envs including R5 and R5X4. R5 and dual-R5 HIV-1 substantially infected BiFC-negative fraction rather than BiFC-positive fraction, indicating the preferential recognition of monomeric CCR5 by R5 and dual-R5 Envs. Although CCR5 antagonists enhanced oligomerization of CCR5, MVC-resistant HIV-1 was found to still recognize both MVC-bound and -unbound forms of monomeric CCR5, suggesting the constrained use of monomeric CCR5 by R5 HIV-1., (Copyright © 2013 Elsevier Inc. All rights reserved.)

A CXCR4 inhibitor-resistant HIV-1 was isolated from a dual-X4 HIV-1 in vitro. The resistant variant displayed competitive resistance to the CXCR4 inhibitor AMD3100, indicating that the resistant variant had a higher affinity for CXCR4 than that of the wild-type HIV-1. Amino acid sequence analyses revealed that the resistant variant harbored amino acid substitutions in the V2, C2, and C4 regions, but no remarkable changes in the V3 loop. Site-directed mutagenesis confirmed that the changes in the C2 and C4 regions were principally involved in the reduced sensitivity to AMD3100. Furthermore, the change in the C4 region was associated with increased sensitivity to soluble CD4, and profoundly enhanced the entry efficiency of the virus. Therefore, it is likely that the resistant variant acquired the higher affinity for CD4/CXCR4 by the changes in non-V3 regions. Taken together, a CXCR4 inhibitor-resistant HIV-1 can evolve using a non-V3 pathway.

Little is known about whether human CD4 positive T cells, the principal natural target of HIV-1, have intrinsic factors, other than the receptor/coreceptor molecules, which modulate the entry efficiency of HIV-1. In the present study, we found that human T cell lines, HUT78 and PM1, were less permissive to VSV-G-mediated HIV-1 infection compared with the Jurkat cell line. Furthermore, HUT78 cells were also less sensitive to HIV-1 Env-mediated infection, while PM1 cells became susceptible to HIV-1. Real-time PCR analyses showed that less susceptibility of the cells to HIV-1 was due to block at, or prior to, reverse transcription of viral RNA. To clarify the entry efficiency of HIV-1 into these cell lines, we analyzed the internalization of p24 Ag into the cytosolic and vesicular fractions of post-nuclear extracts at 4h post-infection. When the cells were infected with HIV-1 pseudotyped with VSV-G, the amount of p24 Ag in the cytosolic fractions in both HUT78 and PM1 cells was lower than that observed in Jurkat cells. In the case of HIV-1 Env-mediated infection, however, PM1 cells exhibited comparable amounts of p24 Ag in the cytosolic fraction compared with Jurkat cells, while the amount of p24 Ag in HUT78 cells remained low. Heterokaryon experiments between susceptible and less susceptible cell lines suggested that some inhibitory factors counteracted VSV-G-mediated viral entry in PM1 and HUT78 cells, and HIV-1 Env-mediated viral entry in HUT78 cells., (Copyright © 2010 Elsevier B.V. All rights reserved.)