RESUMO
Changes to our environment have led to the emergence of human pathogens such as chikungunya virus. Chikungunya virus infection is today a major public health concern. It is a debilitating chronic disease impeding patients' mobility, affecting millions of people. Disease development relies on skeletal muscle infection. The importance of skeletal muscle in chikungunya virus infection led to the hypothesis that it could serve as a viral reservoir and could participate to virus persistence. Here we questioned the interconnection between skeletal muscle cells metabolism, their differentiation stage and the infectivity of the chikungunya virus. We infected human skeletal muscle stem cells at different stages of differentiation with chikungunya virus to study the impact of their metabolism on virus production and inversely the impact of virus on cell metabolism. We observed that chikungunya virus infectivity is cell differentiation and metabolism-dependent. Chikungunya virus interferes with the cellular metabolism in quiescent undifferentiated and proliferative muscle cells. Moreover, activation of chikungunya infected quiescent muscle stem cells, induces their proliferation, increases glycolysis and amplifies virus production. Therefore, our results showed that Chikungunya virus infectivity and the antiviral response of skeletal muscle cells relies on their energetic metabolism and their differentiation stage. Then, muscle stem cells could serve as viral reservoir producing virus after their activation.
Assuntos
Febre de Chikungunya , Vírus Chikungunya , Humanos , Músculo Esquelético/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Diferenciação Celular , Replicação Viral/fisiologiaRESUMO
Study of the human antibody (Ab) response to Aedes salivary proteins can provide new biomarkers to evaluate human exposure to vector bites. The identification of genus- and/or species-specific proteins is necessary to improve the accuracy of biomarkers. We analysed Aedes albopictus immunogenic salivary proteins by 2D immunoproteomic technology and compared the profiles according to human individual exposure to Ae. albopictus or Ae. aegypti bites. Strong antigenicity to Ae. albopictus salivary proteins was detected in all individuals whatever the nature of Aedes exposure. Amongst these antigenic proteins, 68% are involved in blood feeding, including D7 protein family, adenosine deaminase, serpin and apyrase. This study provides an insight into the repertoire of Ae. albopictus immunogenic salivary proteins for the first time.
Assuntos
Aedes/imunologia , Proteínas de Insetos/análise , Proteínas e Peptídeos Salivares/análise , Animais , Feminino , Humanos , Proteínas de Insetos/imunologia , Proteínas e Peptídeos Salivares/imunologiaRESUMO
Increasing evidence suggests that the HIV envelope binds through its surface (SU) gp120 not only to receptors and coreceptors, but also to other components of the cellular membrane where the glycolipids appear to be good candidates. To assess the ability of HIV-1 SU gp120 to penetrate into phospholipid membranes, we carried out a study of the interactions between a recombinant SU gp120 from HIV-1/HXB2 and artificial lipid monolayers mimicking the composition of the outer leaflet of the lymphocytes and which were spread at the air-water interface. We show that the protein, in its aggregated form, has amphipathic properties and that the insertion of this amphipathic species into lipids is favored by the presence of sphingomyelin. Furthermore, cholesterol enhances the penetration into mixed phosphatidylcholine-sphingomyelin monolayers. Coexistence of different physical states of the lipids and thus of domains appears to play a major role for protein penetration independently of the presence of receptors and coreceptors.
Assuntos
Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/metabolismo , HIV-1/patogenicidade , Lipídeos de Membrana/metabolismo , Adsorção , Animais , Bovinos , Membrana Celular/química , Membrana Celular/metabolismo , Membrana Celular/virologia , Colesterol/metabolismo , Gangliosídeo G(M3)/metabolismo , HIV-1/fisiologia , Humanos , Técnicas In Vitro , Lipídeos de Membrana/química , Membranas Artificiais , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismoRESUMO
To evaluate conserved structures of the surface gp120 subunit (SU) of the human immunodeficiency virus type 1 (HIV-1) envelope in gp120-cell interactions, we designed and produced an HIV-1 IIIB (HXB2R) gp120 carrying a deletion of amino acids E61 to S85. This sequence corresponds to a highly conserved predicted amphipathic alpha-helical structure located in the gp120 C1 region. The resultant soluble mutant with a deleted alpha helix 1 (gp120 DeltaalphaHX1) exhibited a strong interaction with CXCR4, although CD4 binding was undetectable. The former interaction was specific since it inhibited the binding of the anti-CXCR4 monoclonal antibody (12G5), as well as SDF1alpha, the natural ligand of CXCR4. Additionally, the mutant gp120 was able to bind to CXCR4(+)/CD4(-) cells but not to CXCR4(-)/CD4(-) cells. Although efficiently expressed on cell surface, HIV envelope harboring the deleted gp120 DeltaalphaHX1 associated with wild-type transmembrane gp41 was unable to induce cell-to-cell fusion with HeLa CD4(+) cells. Nevertheless, the soluble gp120 DeltaalphaHX1 efficiently inhibited a single round of HIV-1 LAI infection in HeLa P4 cells, with a 50% inhibitory concentration of 100 nM. Our data demonstrate that interaction with the CXCR4 coreceptor was maintained in a SUgp120 HIV envelope lacking alphaHX1. Moreover, in the absence of CD4 binding, the interaction of gp120 DeltaalphaHX1 with CXCR4 was sufficient to inhibit HIV-1 infection.
Assuntos
Antígenos CD4/metabolismo , Sequência Conservada , Proteína gp120 do Envelope de HIV/genética , Proteína gp120 do Envelope de HIV/metabolismo , HIV-1/metabolismo , Receptores CXCR4/metabolismo , Sequência de Aminoácidos , Animais , Sítios de Ligação , Células CHO , Membrana Celular/metabolismo , Cricetinae , Expressão Gênica , HIV-1/genética , Células HeLa , Humanos , Dados de Sequência Molecular , Solubilidade , Células Tumorais CultivadasRESUMO
The gp120 envelope glycoprotein of human immunodeficiency virus-1 (HIV-1) interacts with the CXCR4 chemokine receptor, but it is not known whether gp120 activates CXCR4-mediated signaling cascades in the same manner as its natural ligand, SDF1alpha. We assessed the effects of wild-type gp120 and a mutant gp120 that interacts with CXCR4 but not CD4 on CD4(-)/CXCR4(+) cells and CD4(+)/CXCR4(+) cells, respectively. Under both experimental conditions, the interaction of CXCR4 and gp120 resulted in their CD4-independent cointernalization. Both molecules were translocated into early endosomes, whereas neither protein could be detected in late endosomes. Binding of gp120 to CXCR4 resulted in a CD4-independent phosphorylation of Pyk2 and an induction of chemotactic activity, demonstrating that this interaction has functional consequences. Interestingly, however, whereas SDF1alpha activated the ERK/MAP kinase pathway, this cascade was not induced by gp120. Together, these results suggest that the pathology of HIV-1 infection may be modulated by the distinct signal transduction pathway mediated by gp120 upon its interaction with CXCR4.
Assuntos
Antígenos CD/fisiologia , Antígenos CD4/fisiologia , Proteína gp120 do Envelope de HIV/metabolismo , HIV-1/fisiologia , Receptores CXCR4/fisiologia , Linfócitos T/fisiologia , Membrana Celular/fisiologia , Membrana Celular/ultraestrutura , Membrana Celular/virologia , Quimiotaxia de Leucócito , Endossomos/fisiologia , Endossomos/virologia , Citometria de Fluxo , Humanos , Células Jurkat , Transdução de Sinais , Linfócitos T/imunologia , Linfócitos T/virologiaRESUMO
It has been previously shown that the HIV-1 envelope glycoprotein 120 (gp120) activates cell signaling by CXCR4, independently of CD4. The present study examines the involvement of different intracellular signaling pathways and their physiopathologic consequences following the CD4-independent interaction between CXCR4 or CCR5 and gp120 in different cell types: primary T cells, CD4(-)/CXCR4(+)/CCR5(+) T cells, or glioma cells. These interactions were compared with those obtained with natural ligands, stromal cell-derived factor 1 alpha (SDF-1alpha) (CXCL12) and macrophage inflammatory protein 1 beta (MIP-1beta) (CCL4) of their respective coreceptors. Thus, both p38 and SAPK/Jun N-terminal kinase mitogen-activated protein kinases (MAPKs) are activated on stimulation of these cells with either T- or M-tropic gp120, as well as with SDF-1alpha or MIP-1beta. In contrast, extracellular signal-related kinase 1 and 2 MAPKs are only activated by MIP-1beta but not by M-tropic gp120. Importantly, T- and M-tropic gp120 are able to induce the secretion of matrix metalloproteinase 9 (MMP-9), an extracellular metalloproteinase present in cerebrospinal fluid of patients with HIV-1 by T cells or glioma cells. Specific inhibition of MAPK p38 activation resulted in a complete abrogation of the induction of the MMP-9 pathogenic factor expression by gp120 or chemokines in both cell types. Because neurodegenerative features in acquired immune deficiency syndrome dementia may involve demyelinization by MMP-9, the specific targeting of p38 could provide a novel means to control HIV-induced cytopathogenic effects and cell homing to viral replication sites. (Blood. 2001;98:541-547)
Assuntos
Proteína gp120 do Envelope de HIV/farmacologia , Metaloproteinase 9 da Matriz/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Quimiocina CCL3 , Quimiocina CCL4 , Quimiocina CXCL12 , Quimiocinas CXC/metabolismo , Efeito Citopatogênico Viral/efeitos dos fármacos , Efeito Citopatogênico Viral/fisiologia , Ativação Enzimática/efeitos dos fármacos , Humanos , Ligantes , Proteínas Inflamatórias de Macrófagos/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Proteína Quinase 8 Ativada por Mitógeno , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases Ativadas por Mitógeno/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/fisiologia , Receptores CCR5/metabolismo , Receptores CXCR4/metabolismo , Células Tumorais Cultivadas , Proteínas Quinases p38 Ativadas por MitógenoRESUMO
Human apolipoprotein H (apo H) was found to bind specifically to hepatitis B surface antigen (HBsAg) from hepatitis B virus (HBV)-infected individuals. We used recombinant HBsAg proteins to analyze HBV domains recognized by apo H. We showed that the myristylated pre-S1 domain of HBsAg strongly interacted with apo H. This binding involved phospholipid components of the HBV envelope because their removal by detergent prevented apo H-HBsAg interaction. The opposite effects of iron and zinc metal ions on binding suggest that the oxidation of phospholipids also affects apo H-HBsAg interaction. After fractionation of viral particles on a sucrose gradient, and their addition to microtiter plates coated with apo H or anti-HBsAg, we observed that the maximal anti-HBsAg capture activity corresponded to a sucrose concentration of 36%, whereas the maximal apo H capture activity corresponded to a concentration of 39%. Electron microscopy and polymerase chain reaction (PCR) Southern blot studies of these fractions showed that the fraction with maximal apo H binding predominantly contained full Dane particles. Finally, we studied apo H-HBsAg binding relative to the presence of hepatitis B virus markers and observed that apo H binding activity for HBsAg was higher in sera from patients in the active virus replication phase.