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1.
Viruses ; 13(7)2021 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-34372572

RESUMO

Human APOBEC3 (apolipoprotein B mRNA-editing catalytic polypeptide-like 3) enzymes are capable of inhibiting a wide range of endogenous and exogenous viruses using deaminase and deaminase-independent mechanisms. These enzymes are essential components of our innate immune system, as evidenced by (a) their strong positive selection and expansion in primates, (b) the evolution of viral counter-defense mechanisms, such as proteasomal degradation mediated by HIV Vif, and (c) hypermutation and inactivation of a large number of integrated HIV-1 proviruses. Numerous APOBEC3 single nucleotide polymorphisms, haplotypes, and splice variants have been identified in humans. Several of these variants have been reported to be associated with differential antiviral immunity. This review focuses on the current knowledge in the field about these natural variations and their roles in infectious diseases.


Assuntos
Desaminases APOBEC/genética , Desaminases APOBEC/metabolismo , Viroses/genética , Citidina Desaminase/genética , Citosina Desaminase/genética , HIV-1/fisiologia , Vírus da Hepatite B/fisiologia , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata/imunologia , Polimorfismo Genético/genética , Isoformas de Proteínas/genética , Viroses/metabolismo , Replicação Viral/genética
2.
Science ; 373(6555): 700-704, 2021 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-34353956

RESUMO

Gag, the primary structural protein of HIV-1, is recruited to the plasma membrane for virus assembly by its matrix (MA) domain. Gag is subsequently cleaved into its component domains, causing structural maturation to repurpose the virion for cell entry. We determined the structure and arrangement of MA within immature and mature HIV-1 through cryo-electron tomography. We found that MA rearranges between two different hexameric lattices upon maturation. In mature HIV-1, a lipid extends out of the membrane to bind with a pocket in MA. Our data suggest that proteolytic maturation of HIV-1 not only assembles the viral capsid surrounding the genome but also repurposes the membrane-bound MA lattice for an entry or postentry function and results in the partial removal of up to 2500 lipids from the viral membrane.


Assuntos
Antígenos HIV/química , Antígenos HIV/metabolismo , HIV-1/química , HIV-1/fisiologia , Envelope Viral/metabolismo , Produtos do Gene gag do Vírus da Imunodeficiência Humana/química , Produtos do Gene gag do Vírus da Imunodeficiência Humana/metabolismo , Capsídeo/química , Capsídeo/fisiologia , Tomografia com Microscopia Eletrônica , HIV-1/ultraestrutura , Bicamadas Lipídicas , Lipídeos de Membrana/metabolismo , Modelos Moleculares , Conformação Proteica , Domínios Proteicos , Estrutura Secundária de Proteína , Envelope Viral/química , Envelope Viral/ultraestrutura , Vírion/química , Vírion/fisiologia , Vírion/ultraestrutura , Montagem de Vírus , Produtos do Gene env do Vírus da Imunodeficiência Humana/química , Produtos do Gene env do Vírus da Imunodeficiência Humana/metabolismo
3.
J Immunol ; 207(5): 1239-1249, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34389623

RESUMO

HIV-1 infection substantially increases the risk of developing tuberculosis (TB). Mechanisms such as defects in the Th1 response to Mycobacterium tuberculosis in HIV-infected persons have been widely reported. However, Th1-independent mechanisms also contribute to protection against TB. To identify a broader spectrum of defects in TB immunity during HIV infection, we examined IL-17A and IL-22 production in response to mycobacterial Ags in peripheral blood of persons with latent TB infection and HIV coinfection. Upon stimulating with mycobacterial Ags, we observed a distinct CD4+ Th lineage producing IL-22 in the absence of IL-17A and IFN-γ. Mycobacteria-specific Th22 cells were present at high frequencies in blood and contributed up to 50% to the CD4+ T cell response to mycobacteria, comparable in magnitude to the IFN-γ Th1 response (median 0.91% and 0.55%, respectively). Phenotypic characterization of Th22 cells revealed that their memory differentiation was similar to M. tuberculosis-specific Th1 cells (i.e., predominantly early differentiated CD45RO+CD27+ phenotype). Moreover, CCR6 and CXCR3 expression profiles of Th22 cells were similar to Th17 cells, whereas their CCR4 and CCR10 expression patterns displayed an intermediate phenotype between Th1 and Th17 cells. Strikingly, mycobacterial IL-22 responses were 3-fold lower in HIV-infected persons compared with uninfected persons, and the magnitude of responses correlated inversely with HIV viral load. These data provide important insights into mycobacteria-specific Th subsets in humans and suggest a potential role for IL-22 in protection against TB during HIV infection. Further studies are needed to fully elucidate the role of IL-22 in protective TB immunity.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/imunologia , HIV-1/fisiologia , Interleucinas/metabolismo , Tuberculose Latente/imunologia , Mycobacterium tuberculosis/fisiologia , Subpopulações de Linfócitos T/imunologia , Adulto , Células Cultivadas , Coinfecção , Feminino , Soropositividade para HIV , Humanos , Interleucina-17/metabolismo , Masculino , África do Sul , Carga Viral , Adulto Jovem
4.
Top Antivir Med ; 29(3): 355-360, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34370417

RESUMO

The Conference on Retroviruses and Opportunistic Infections (CROI) serves as one of the most highly visible platforms upon which researchers gather to share the most recent findings on HIV/AIDS and, recently, on SARS-CoV-2 research. Research presentations on the novel coronavirus SARS-CoV-2 have become an increasing fixture at the conference since it was first covered at last year's conference. Although CROI 2021 was virtual, the organizers coordinated a seamless platform for presentations and poster sessions that effectively engaged the audience. CROI 2021 had a strong showing in terms of basic science presentations on HIV-1 and on SARS-CoV-2. Highlights included new insights into some of the more elusive steps in the viral replication cycle as well as new findings on immune escape strategies employed by SARS-CoV-2. The new investigator workshop has become a valuable resource that can be used by early stage and established investigators alike to receive state-of-the-art updates on research areas that might be outside their immediate areas of research. The new investigator workshop featured engaging presentations on novel aspects of HIV-1 and SARS-CoV-2 replication, impact of host immunity on HIV-1 and SARS-CoV-2, and approaches to assessing viral reservoir dynamics and strategies for viral reservoir elimination.


Assuntos
Pesquisa Biomédica , Congressos como Assunto , HIV-1/imunologia , SARS-CoV-2/imunologia , HIV-1/fisiologia , Humanos , SARS-CoV-2/fisiologia , Latência Viral
5.
BMC Infect Dis ; 21(1): 655, 2021 Jul 07.
Artigo em Inglês | MEDLINE | ID: mdl-34233649

RESUMO

BACKGROUND: Macrophages, besides resting latently infected CD4+ T cells, constitute the predominant stable, major non-T cell HIV reservoirs. Therefore, it is essential to eliminate both latently infected CD4+ T cells and tissue macrophages to completely eradicate HIV in patients. Until now, most of the research focus is directed towards eliminating latently infected CD4+ T cells. However, few approaches have been directed at killing of HIV-infected macrophages either in vitro or in vivo. HIV infection dysregulates the expression of many host genes essential for the survival of infected cells. We postulated that exploiting this alteration may yield novel targets for the selective killing of infected macrophages. METHODS: We applied a pooled shRNA-based genome-wide approach by employing a lentivirus-based library of shRNAs to screen novel gene targets whose inhibition should selectively induce apoptosis in HIV-infected macrophages. Primary human MDMs were infected with HIV-eGFP and HIV-HSA viruses. Infected MDMs were transfected with siRNAs specific for the promising genes followed by analysis of apoptosis by flow cytometry using labelled Annexin-V in HIV-infected, HIV-exposed but uninfected bystander MDMs and uninfected MDMs. The results were analyzed using student's t-test from at least four independent experiments. RESULTS: We validated 28 top hits in two independent HIV infection models. This culminated in the identification of four target genes, Cox7a2, Znf484, Cstf2t, and Cdk2, whose loss-of-function induced apoptosis preferentially in HIV-infected macrophages. Silencing these single genes killed significantly higher number of HIV-HSA-infected MDMs compared to the HIV-HSA-exposed, uninfected bystander macrophages, indicating the specificity in the killing of HIV-infected macrophages. The mechanism governing Cox7a2-mediated apoptosis of HIV-infected macrophages revealed that targeting respiratory chain complex II and IV genes also selectively induced apoptosis of HIV-infected macrophages possibly through enhanced ROS production. CONCLUSIONS: We have identified above-mentioned novel genes and specifically the respiratory chain complex II and IV genes whose silencing may cause selective elimination of HIV-infected macrophages and eventually the HIV-macrophage reservoirs. The results highlight the potential of the identified genes as targets for eliminating HIV-infected macrophages in physiological environment as part of an HIV cure strategy.


Assuntos
Apoptose/genética , Proteínas de Fluorescência Verde , Infecções por HIV , Macrófagos , RNA Interferente Pequeno , Linfócitos T CD4-Positivos/virologia , Estudo de Associação Genômica Ampla , Infecções por HIV/genética , Infecções por HIV/virologia , HIV-1/fisiologia , Humanos , Linfócitos T
6.
Int J Mol Sci ; 22(13)2021 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-34206330

RESUMO

The 'shock-and-kill' strategy to purge the latent HIV reservoir relies on latency-reversing agents (LRAs) to reactivate the provirus and subsequent immune-mediated killing of HIV-expressing cells. Yet, clinical trials employing histone deacetylase inhibitors (HDACis; Vorinostat, Romidepsin, Panobinostat) as LRAs failed to reduce the HIV reservoir size, stressing the need for more effective latency reversal strategies, such as 2-LRA combinations, and enhancement of the immune responses. Interestingly, several LRAs are employed to treat cancer because they up-modulate ligands for the NKG2D NK-cell activating receptor on tumor cells. Therefore, using in vitro T cell models of HIV latency and NK cells, we investigated the capacity of HDACis, either alone or combined with a distinct LRA, to potentiate the NKG2D/NKG2D ligands axis. While Bortezomib proteasome inhibitor was toxic for both T and NK cells, the GS-9620 TLR-7 agonist antagonized HIV reactivation and NKG2D ligand expression by HDACis. Conversely, co-administration of the Prostratin PKC agonist attenuated HDACi toxicity and, when combined with Romidepsin, stimulated HIV reactivation and further up-modulated NKG2D ligands on HIV+ T cells and NKG2D on NK cells, ultimately boosting NKG2D-mediated viral suppression by NK cells. These findings disclose limitations of LRA candidates and provide evidence that NK cell suppression of reactivated HIV may be modulated by specific 2-LRA combinations.


Assuntos
Infecções por HIV/tratamento farmacológico , HIV-1/fisiologia , Inibidores de Histona Desacetilases/uso terapêutico , Células Matadoras Naturais/imunologia , Linfócitos T/virologia , Latência Viral , Infecções por HIV/imunologia , Infecções por HIV/fisiopatologia , Infecções por HIV/terapia , Humanos , Células Matadoras Naturais/fisiologia
7.
Lancet HIV ; 8(7): e397-e407, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34197772

RESUMO

BACKGROUND: In DISCOVER, a multinational, randomised controlled trial, emtricitabine and tenofovir alafenamide compared with emtricitabine and tenofovir disoproxil fumarate showed non-inferior efficacy for HIV prevention and improved bone mineral density and renal safety biomarkers at week 48. We report outcomes analysed after all participants had completed 96 weeks of follow-up. METHODS: This study is an ongoing, randomised, double-blind, multicentre, active-controlled, phase 3, non-inferiority trial done at 94 community, public health, and hospital-associated clinics located in Europe and North America. Adult cisgender men and transgender women who have sex with men, both with a high risk of acquiring HIV as determined by self-reported sexual behaviour or recent sexually transmitted infections, were randomly assigned (1:1) to receive either emtricitabine and tenofovir alafenamide (200/25 mg) tablets daily, with matched placebo tablets (emtricitabine and tenofovir alafenamide group), or emtricitabine and tenofovir disoproxil fumarate (200/300 mg) tablets daily, with matched placebo tablets (emtricitabine and tenofovir disoproxil fumarate group). The primary efficacy outcome was incident HIV infection. Incidence of HIV-1 infection per 100 person-years was assessed when the last participant had completed 96 weeks of follow-up. This trial is registered with ClinicalTrials.gov, number NCT02842086. FINDINGS: Between Sept 13, 2016, and June 30, 2017, 5387 participants were randomly assigned to receive emtricitabine and tenofovir alafenamide (n=2694) or emtricitabine and tenofovir disoproxil fumarate (n=2693), contributing 10 081 person-years of follow-up. At 96 weeks of follow-up, there were eight HIV infections in participants who had received emtricitabine and tenofovir alafenamide (0·16 infections per 100 person-years [95% CI 0·07-0·31]) and 15 in participants who had received emtricitabine and tenofovir disoproxil fumarate (0·30 infections per 100 person-years [0·17-0·49]). Emtricitabine and tenofovir alafenamide maintained its non-inferiority to emtricitabine and tenofovir disoproxil fumarate for HIV prevention (IRR 0·54 [95% CI 0·23-1·26]). Approximately 78-82% of participants reported taking study medication more than 95% of the time across all study visits. Rates of sexually transmitted infections remained high and similar across groups (21 cases per 100 person-years for rectal gonorrhoea and 28 cases per 100 person-years for rectal chlamydia). Emtricitabine and tenofovir alafenamide continued to show superiority over emtricitabine and tenofovir disoproxil fumarate in all but one of the six prespecified bone mineral density and renal biomarkers. There was more weight gain among participants who had received emtricitabine and tenofovir alafenamide (median weight gain 1·7 kg vs 0·5 kg, p<0·0001). INTERPRETATION: Emtricitabine and tenofovir alafenamide is safe and effective for longer-term pre-exposure prophylaxis in cisgender men and transgender women who have sex with men. FUNDING: Gilead Sciences.


Assuntos
Adenina/análogos & derivados , Alanina/uso terapêutico , Fármacos Anti-HIV/uso terapêutico , Emtricitabina/uso terapêutico , Infecções por HIV/prevenção & controle , Organofosfonatos/uso terapêutico , Tenofovir/uso terapêutico , Adenina/efeitos adversos , Adenina/uso terapêutico , Adulto , Idoso , Alanina/efeitos adversos , Fármacos Anti-HIV/efeitos adversos , Método Duplo-Cego , Quimioterapia Combinada/efeitos adversos , Emtricitabina/efeitos adversos , Feminino , Seguimentos , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Humanos , Masculino , Pessoa de Meia-Idade , Organofosfonatos/efeitos adversos , Profilaxia Pré-Exposição , Tenofovir/efeitos adversos , Resultado do Tratamento , Adulto Jovem
8.
Nat Commun ; 12(1): 4582, 2021 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-34321470

RESUMO

SAMHD1 is a cellular triphosphohydrolase (dNTPase) proposed to inhibit HIV-1 reverse transcription in non-cycling immune cells by limiting the supply of the dNTP substrates. Yet, phosphorylation of T592 downregulates SAMHD1 antiviral activity, but not its dNTPase function, implying that additional mechanisms contribute to viral restriction. Here, we show that SAMHD1 is SUMOylated on residue K595, a modification that relies on the presence of a proximal SUMO-interacting motif (SIM). Loss of K595 SUMOylation suppresses the restriction activity of SAMHD1, even in the context of the constitutively active phospho-ablative T592A mutant but has no impact on dNTP depletion. Conversely, the artificial fusion of SUMO2 to a non-SUMOylatable inactive SAMHD1 variant restores its antiviral function, a phenotype that is reversed by the phosphomimetic T592E mutation. Collectively, our observations clearly establish that lack of T592 phosphorylation cannot fully account for the restriction activity of SAMHD1. We find that SUMOylation of K595 is required to stimulate a dNTPase-independent antiviral activity in non-cycling immune cells, an effect that is antagonized by cyclin/CDK-dependent phosphorylation of T592 in cycling cells.


Assuntos
Ciclo Celular/fisiologia , HIV-1/fisiologia , Proteína 1 com Domínio SAM e Domínio HD/genética , Proteína 1 com Domínio SAM e Domínio HD/metabolismo , Sumoilação/fisiologia , Substituição de Aminoácidos , Células HEK293 , Infecções por HIV/virologia , Humanos , Lisina , Mutação , Fosforilação , Proteína 1 com Domínio SAM e Domínio HD/química , Células U937
9.
Nat Commun ; 12(1): 4503, 2021 07 23.
Artigo em Inglês | MEDLINE | ID: mdl-34301927

RESUMO

Promoter-proximal pausing of RNA polymerase II is a key process regulating gene expression. In latent HIV-1 cells, it prevents viral transcription and is essential for latency maintenance, while in acutely infected cells the viral factor Tat releases paused polymerase to induce viral expression. Pausing is fundamental for HIV-1, but how it contributes to bursting and stochastic viral reactivation is unclear. Here, we performed single molecule imaging of HIV-1 transcription. We developed a quantitative analysis method that manages multiple time scales from seconds to days and that rapidly fits many models of promoter dynamics. We found that RNA polymerases enter a long-lived pause at latent HIV-1 promoters (>20 minutes), thereby effectively limiting viral transcription. Surprisingly and in contrast to current models, pausing appears stochastic and not obligatory, with only a small fraction of the polymerases undergoing long-lived pausing in absence of Tat. One consequence of stochastic pausing is that HIV-1 transcription occurs in bursts in latent cells, thereby facilitating latency exit and providing a rationale for the stochasticity of viral rebounds.


Assuntos
Regulação Viral da Expressão Gênica , Infecções por HIV/genética , HIV-1/genética , Regiões Promotoras Genéticas/genética , Latência Viral/genética , Algoritmos , RNA Polimerases Dirigidas por DNA/metabolismo , Infecções por HIV/metabolismo , Infecções por HIV/virologia , HIV-1/fisiologia , Células HeLa , Humanos , Modelos Genéticos , Processos Estocásticos , Fatores de Tempo , Ativação Viral/genética , Produtos do Gene tat do Vírus da Imunodeficiência Humana/genética
10.
Viruses ; 13(6)2021 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-34200141

RESUMO

The transmission of viruses from animal hosts into humans have led to the emergence of several diseases. Usually these cross-species transmissions are blocked by host restriction factors, which are proteins that can block virus replication at a specific step. In the natural virus host, the restriction factor activity is usually suppressed by a viral antagonist protein, but this is not the case for restriction factors from an unnatural host. However, due to ongoing viral evolution, sometimes the viral antagonist can evolve to suppress restriction factors in a new host, enabling cross-species transmission. Here we examine the classical case of this paradigm by reviewing research on APOBEC3 restriction factors and how they can suppress human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV). APOBEC3 enzymes are single-stranded DNA cytidine deaminases that can induce mutagenesis of proviral DNA by catalyzing the conversion of cytidine to promutagenic uridine on single-stranded viral (-)DNA if they escape the HIV/SIV antagonist protein, Vif. APOBEC3 degradation is induced by Vif through the proteasome pathway. SIV has been transmitted between Old World Monkeys and to hominids. Here we examine the adaptations that enabled such events and the ongoing impact of the APOBEC3-Vif interface on HIV in humans.


Assuntos
Desaminases APOBEC/genética , Interações Hospedeiro-Patógeno/genética , Infecções por Lentivirus/genética , Infecções por Lentivirus/transmissão , Lentivirus de Primatas/fisiologia , Zoonoses Virais/transmissão , Animais , Produtos do Gene vif/química , Produtos do Gene vif/metabolismo , Infecções por HIV/genética , Infecções por HIV/transmissão , Infecções por HIV/virologia , HIV-1/fisiologia , Humanos , Infecções por Lentivirus/virologia , Ligação Proteica , Isoformas de Proteínas , Relação Estrutura-Atividade , Produtos do Gene vif do Vírus da Imunodeficiência Humana/química , Produtos do Gene vif do Vírus da Imunodeficiência Humana/metabolismo
11.
Viruses ; 13(6)2021 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-34201134

RESUMO

To explore the sequence context-dependent nature of the human immunodeficiency virus type 1 (HIV-1) protease's specificity and to provide a rationale for viral mutagenesis to study the potential role of the nucleocapsid (NC) processing in HIV-1 replication, synthetic oligopeptide substrates representing the wild-type and modified versions of the proximal cleavage site of HIV-1 NC were assayed as substrates of the HIV-1 protease (PR). The S1' substrate binding site of HIV-1 PR was studied by an in vitro assay using KIVKCF↓NCGK decapeptides having amino acid substitutions of N17 residue of the cleavage site of the first zinc-finger domain, and in silico calculations were also performed to investigate amino acid preferences of S1' site. Second site substitutions have also been designed to produce "revertant" substrates and convert a non-hydrolysable sequence (having glycine in place of N17) to a substrate. The specificity constants obtained for peptides containing non-charged P1' substitutions correlated well with the residue volume, while the correlation with the calculated interaction energies showed the importance of hydrophobicity: interaction energies with polar residues were related to substantially lower specificity constants. Cleavable "revertants" showed one residue shift of cleavage position due to an alternative productive binding mode, and surprisingly, a double cleavage of a substrate was also observed. The results revealed the importance of alternative binding possibilities of substrates into the HIV-1 PR. The introduction of the "revertant" mutations into infectious virus clones may provide further insights into the potential role of NC processing in the early phase of the viral life-cycle.


Assuntos
Infecções por HIV/virologia , Protease de HIV/metabolismo , HIV-1/fisiologia , Proteínas do Nucleocapsídeo/metabolismo , Dedos de Zinco , Sequência de Aminoácidos , Sítios de Ligação , Desenho de Fármacos , Protease de HIV/química , Humanos , Modelos Moleculares , Proteínas do Nucleocapsídeo/química , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Proteólise , Proteínas Recombinantes , Relação Estrutura-Atividade , Especificidade por Substrato
12.
Commun Biol ; 4(1): 861, 2021 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-34253821

RESUMO

Mucosal exposure to infected semen accounts for the majority of HIV-1 transmission events, with rectal intercourse being the route with the highest estimated risk of transmission. Yet, the impact of semen inflammation on colorectal HIV-1 transmission has never been addressed. Here we use cynomolgus macaques colorectal tissue explants to explore the effect of leukocytospermia, indicative of male genital tract inflammation, on SIVmac251 infection. We show that leukocytospermic seminal plasma (LSP) has significantly higher concentration of a number of pro-inflammatory molecules compared to normal seminal plasma (NSP). In virus-exposed explants, LSP enhance SIV infection more efficiently than NSP, being the increased viral replication linked to the level of inflammatory and immunomodulatory cytokines. Moreover, LSP induce leukocyte accumulation on the apical side of the colorectal lamina propria and the recruitment of a higher number of intraepithelial dendritic cells than with NSP. These results suggest that the outcome of mucosal HIV-1 infection is influenced by the inflammatory state of the semen donor, and provide further insights into mucosal SIV/HIV-1 pathogenesis.


Assuntos
Colo/virologia , Células Dendríticas/virologia , Reto/virologia , Sêmen/virologia , Vírus da Imunodeficiência Símia/fisiologia , Replicação Viral/fisiologia , Animais , Colo/metabolismo , Citocinas/metabolismo , Infecções por HIV/metabolismo , Infecções por HIV/transmissão , Infecções por HIV/virologia , HIV-1/fisiologia , Leucócitos/metabolismo , Leucócitos/patologia , Leucócitos/virologia , Macaca mulatta , Masculino , Reto/metabolismo , Síndrome de Imunodeficiência Adquirida dos Símios/metabolismo , Síndrome de Imunodeficiência Adquirida dos Símios/transmissão , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Técnicas de Cultura de Tecidos
13.
Nat Microbiol ; 6(8): 1031-1042, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34282309

RESUMO

The antiviral cytokine interferon activates expression of interferon-stimulated genes to establish an antiviral state. Myxovirus resistance 2 (MX2, also known as MxB) is an interferon-stimulated gene that inhibits the nuclear import of HIV-1 and interacts with the viral capsid and cellular nuclear transport machinery. Here, we identified the myosin light chain phosphatase (MLCP) subunits myosin phosphatase target subunit 1 (MYPT1) and protein phosphatase 1 catalytic subunit-ß (PPP1CB) as positively-acting regulators of MX2, interacting with its amino-terminal domain. We demonstrated that serine phosphorylation of the N-terminal domain at positions 14, 17 and 18 suppresses MX2 antiviral function, prevents interactions with the HIV-1 capsid and nuclear transport factors, and is reversed by MLCP. Notably, serine phosphorylation of the N-terminal domain also impedes MX2-mediated inhibition of nuclear import of cellular karyophilic cargo. We also found that interferon treatment reduces levels of phosphorylation at these serine residues and outline a homeostatic regulatory mechanism in which repression of MX2 by phosphorylation, together with MLCP-mediated dephosphorylation, balances the deleterious effects of MX2 on normal cell function with innate immunity against HIV-1.


Assuntos
Infecções por HIV/imunologia , HIV-1/imunologia , Imunidade Inata , Proteínas de Resistência a Myxovirus/química , Proteínas de Resistência a Myxovirus/imunologia , Motivos de Aminoácidos , Infecções por HIV/genética , Infecções por HIV/virologia , HIV-1/genética , HIV-1/fisiologia , Células HeLa , Humanos , Fosfatase de Miosina-de-Cadeia-Leve/genética , Fosfatase de Miosina-de-Cadeia-Leve/imunologia , Fosfatase de Miosina-de-Cadeia-Leve/metabolismo , Proteínas de Resistência a Myxovirus/genética , Fosforilação , Domínios Proteicos , Proteína Fosfatase 1/genética , Proteína Fosfatase 1/imunologia , Serina/metabolismo
15.
Viruses ; 13(5)2021 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-34066522

RESUMO

Viral entry into host cells is a critical step in the viral life cycle. HIV-1 entry is mediated by the sole surface envelope glycoprotein Env and is initiated by the interaction between Env and the host receptor CD4. This interaction, referred to as the attachment step, has long been considered an attractive target for inhibitor discovery and development. Fostemsavir, recently approved by the FDA, represents the first-in-class drug in the attachment inhibitor class. This review focuses on the discovery of temsavir (the active compound of fostemsavir) and analogs, mechanistic studies that elucidated the mode of action, and structural studies that revealed atomic details of the interaction between HIV-1 Env and attachment inhibitors. Challenges associated with emerging resistance mutations to the attachment inhibitors and the development of next-generation attachment inhibitors are also highlighted.


Assuntos
Fármacos Anti-HIV/farmacologia , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Organofosfatos/farmacologia , Piperazinas/farmacologia , Animais , Fármacos Anti-HIV/química , Infecções por HIV/tratamento farmacológico , HIV-1/genética , Humanos , Organofosfatos/química , Piperazinas/química , Internalização do Vírus/efeitos dos fármacos
17.
Viruses ; 13(5)2021 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-34064404

RESUMO

The HIV replication cycle depends on the interaction of viral proteins with proteins of the host. Unraveling host-pathogen interactions during the infection is of great importance for understanding the pathogenesis and the development of antiviral therapies. To date HIV uncoating and nuclear import are the most debated steps of the HIV-1 replication cycle. Despite numerous studies during past decades, there is still much controversy with respect to the identity and the role of viral and host factors involved in these processes. In this review, we provide a comprehensive overview on the role of transportin-SR2 as a host cell factor during active nuclear transport.


Assuntos
Infecções por HIV/metabolismo , Infecções por HIV/virologia , HIV-1/fisiologia , Interações Hospedeiro-Patógeno , Replicação Viral , beta Carioferinas/metabolismo , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Capsídeo/metabolismo , Ciclofilina A/farmacologia , Integrase de HIV/metabolismo , Humanos , Modelos Biológicos , Ligação Proteica , Fatores de Poliadenilação e Clivagem de mRNA/metabolismo
18.
AAPS PharmSciTech ; 22(5): 171, 2021 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-34100170

RESUMO

Macrophages act as a cellular reservoir in HIV infection. Elimination of HIV from macrophages has been an unfulfilled dream due to the failure of drugs to reach them. To address this, we developed CD44 receptor-targeted, novel hyaluronic acid (HA)-coated nanostructured lipid carriers (NLCs) of efavirenz via washless layer-by-layer (LbL) assembly of HA and polyallylamine hydrochloride (PAH). NLCs were subjected to TEM analysis, size and zeta potential, in vitro release and encapsulation efficiency studies. The uptake of NLCs in THP-1 cells was studied using fluorescence microscopy and flow cytometry. The anti-HIV efficacy was evaluated using p24 antigen inhibition assay. NLCs were found to be spherical in shape with anionic zeta potential (-23.66 ± 0.87 mV) and 241.83 ± 5.38 nm particle size. NLCs exhibited prolonged release of efavirenz during in vitro drug release studies. Flow cytometry revealed 1.73-fold higher uptake of HA-coated NLCs in THP-1 cells. Cytotoxicity studies showed no significant change in cell viability in presence of NLCs as compared with the control. HA-coated NLCs distributed throughout the cell including cytoplasm, plasma membrane and nucleus, as observed during fluorescence microscopy. HA-coated NLCs demonstrated consistent and significantly higher inhibition (81.26 ± 1.70%) of p24 antigen which was 2.08-fold higher than plain NLCs. The obtained results suggested preferential uptake of HA-coated NLCs via CD44-mediated uptake. The present finding demonstrates that HA-based CD44 receptor targeting in HIV infection is an attractive strategy for maximising the drug delivery to macrophages and achieve effective viral inhibition.


Assuntos
Portadores de Fármacos/administração & dosagem , HIV-1/efeitos dos fármacos , Receptores de Hialuronatos , Macrófagos/efeitos dos fármacos , Nanoestruturas/administração & dosagem , Inibidores da Transcriptase Reversa/administração & dosagem , Alcinos/administração & dosagem , Alcinos/síntese química , Alcinos/metabolismo , Benzoxazinas/administração & dosagem , Benzoxazinas/síntese química , Benzoxazinas/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Ciclopropanos/administração & dosagem , Ciclopropanos/síntese química , Ciclopropanos/metabolismo , Relação Dose-Resposta a Droga , Portadores de Fármacos/síntese química , Portadores de Fármacos/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Células HEK293 , Infecções por HIV/tratamento farmacológico , Infecções por HIV/metabolismo , HIV-1/fisiologia , Humanos , Receptores de Hialuronatos/metabolismo , Lipídeos/administração & dosagem , Lipídeos/síntese química , Macrófagos/metabolismo , Nanoestruturas/química , Inibidores da Transcriptase Reversa/síntese química , Inibidores da Transcriptase Reversa/metabolismo , Células THP-1
19.
Viruses ; 13(5)2021 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-33946474

RESUMO

Human immunodeficiency virus (HIV) is associated with neuroendocrine dysfunction which may contribute to co-morbid stress-sensitive disorders. The hypothalamic-pituitary-adrenal (HPA) or -gonadal (HPG) axes are perturbed in up to 50% of HIV patients. The mechanisms are not known, but we have found the HIV-1 trans-activator of transcription (Tat) protein to recapitulate the clinical phenotype in male mice. We hypothesized that HPA and/or HPG dysregulation contributes to Tat-mediated interactions with oxycodone, an opioid often prescribed to HIV patients, in females. Female mice that conditionally-expressed the Tat1-86 protein [Tat(+) mice] or their counterparts that did not [Tat(-) control mice] were exposed to forced swim stress (or not) and behaviorally-assessed for motor and anxiety-like behavior. Some mice had glucocorticoid receptors (GR) or corticotropin-releasing factor receptors (CRF-R) pharmacologically inhibited. Some mice were ovariectomized (OVX). As seen previously in males, Tat elevated basal corticosterone levels and potentiated oxycodone's psychomotor activity in females. Unlike males, females did not demonstrate adrenal insufficiency and oxycodone potentiation was not regulated by GRs or CRF-Rs. Rather OVX attenuated Tat/oxycodone interactions. Either Tat or oxycodone increased anxiety-like behavior and their combination increased hypothalamic allopregnanolone. OVX increased basal hypothalamic allopregnanolone and obviated Tat or oxycodone-mediated fluctuations. Together, these data provide further evidence for Tat-mediated dysregulation of the HPA axis and reveal the importance of HPG axis regulation in females. HPA/HPG disruption may contribute vulnerability to affective and substance use disorders.


Assuntos
Infecções por HIV/complicações , Infecções por HIV/virologia , HIV-1/fisiologia , Sistemas Neurossecretores/metabolismo , Sistemas Neurossecretores/fisiopatologia , Oxicodona/farmacologia , Desempenho Psicomotor/efeitos dos fármacos , Produtos do Gene tat do Vírus da Imunodeficiência Humana/metabolismo , Animais , Comportamento Animal , Modelos Animais de Doenças , Ciclo Estral , Feminino , Humanos , Sistema Hipotálamo-Hipofisário/efeitos dos fármacos , Sistema Hipotálamo-Hipofisário/metabolismo , Camundongos , Atividade Motora , Sistemas Neurossecretores/efeitos dos fármacos , Sistema Hipófise-Suprarrenal/efeitos dos fármacos , Sistema Hipófise-Suprarrenal/metabolismo , Pregnanolona/sangue , Pregnanolona/metabolismo , Esteroides/sangue
20.
Nat Commun ; 12(1): 2743, 2021 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-33980829

RESUMO

INI1/SMARCB1 binds to HIV-1 integrase (IN) through its Rpt1 domain and exhibits multifaceted role in HIV-1 replication. Determining the NMR structure of INI1-Rpt1 and modeling its interaction with the IN-C-terminal domain (IN-CTD) reveal that INI1-Rpt1/IN-CTD interface residues overlap with those required for IN/RNA interaction. Mutational analyses validate our model and indicate that the same IN residues are involved in both INI1 and RNA binding. INI1-Rpt1 and TAR RNA compete with each other for IN binding with similar IC50 values. INI1-interaction-defective IN mutant viruses are impaired for incorporation of INI1 into virions and for particle morphogenesis. Computational modeling of IN-CTD/TAR complex indicates that the TAR interface phosphates overlap with negatively charged surface residues of INI1-Rpt1 in three-dimensional space, suggesting that INI1-Rpt1 domain structurally mimics TAR. This possible mimicry between INI1-Rpt1 and TAR explains the mechanism by which INI1/SMARCB1 influences HIV-1 late events and suggests additional strategies to inhibit HIV-1 replication.


Assuntos
Integrase de HIV/metabolismo , HIV-1/fisiologia , RNA Viral/metabolismo , Proteína SMARCB1/metabolismo , Replicação Viral , Genoma Viral , Integrase de HIV/química , Integrase de HIV/genética , Interações Hospedeiro-Patógeno , Humanos , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Simulação de Acoplamento Molecular , Ligação Proteica , Domínios Proteicos , RNA Viral/química , Proteína SMARCB1/química , Proteína SMARCB1/genética , Vírion/crescimento & desenvolvimento , Vírion/metabolismo
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