RESUMEN
DC-SIGN, a C-type lectin expressed on dendritic cells (DCs), can sequester human immunodeficiency virus (HIV) virions in multivesicular bodies. Here, using large-scale gene expression profiling and tyrosine-phosphorylated proteome analyses, we characterized signaling mediated by DC-SIGN after activation by either HIV or a DC-SIGN-specific antibody. Activation of DC-SIGN resulted in downregulation of genes encoding major histocompatibility complex class II, Jagged 1 and interferon-response molecules and upregulation of the gene encoding transcription factor ATF3. Phosphorylated proteome analysis showed that HIV- or antibody-stimulated DC-SIGN signaling was mediated by the Rho guanine nucleotide-exchange factor LARG and led to increased Rho-GTPase activity. Activation of LARG in DCs exposed to HIV was required for the formation of virus-T cell synapses. Thus, HIV sequestration by and stimulation of DC-SIGN helps HIV evade immune responses and spread to cells.
Asunto(s)
Moléculas de Adhesión Celular/inmunología , Células Dendríticas/enzimología , Células Dendríticas/inmunología , VIH-1/fisiología , Lectinas Tipo C/inmunología , Receptores de Superficie Celular/inmunología , Replicación Viral , Proteínas de Unión al GTP rho/metabolismo , Secuencia de Aminoácidos , Diferenciación Celular , Células Cultivadas , Células Dendríticas/citología , Activación Enzimática , Perfilación de la Expresión Génica , Factores de Intercambio de Guanina Nucleótido/química , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/metabolismo , Humanos , Datos de Secuencia Molecular , Fenotipo , Fosfoserina/metabolismo , Factores de Intercambio de Guanina Nucleótido Rho , Transducción de Señal , Linfocitos T/inmunología , Linfocitos T/metabolismo , Transcripción Genética/genéticaRESUMEN
Lentiviral Nef increases T cell signaling activity, but the molecular nature of the stimulus involved is incompletely described. We explored CD4 T cell lipid raft composition in the presence and absence of Nef. Here, the E2 ubiquitin-conjugating enzyme UbcH7, which acts in conjunction with c-Cbl, is absent from lipid rafts. This Nef-mediated exclusion is associated with failure of ubiquitination of activated Vav. In the presence of Nef, lipid raft Cdc42 is activated and forms a ternary complex between the c-Cbl-interacting protein p85Cool-1/betaPix and c-Cbl, displacing UbcH7 from rafts. Suppression of p85Cool-1/betaPix expression restores UbcH7 raft localization and Vav ubiquitination and diminishes Cdc42 activity. Moreover, p85Cool-1/betaPix knockdown attenuates HIV replication. Thresholds for activation of signaling involve the intricate balance of positive and negative regulators. Here we provide evidence for Nef disruption of a negative regulator of T cell signaling in promoting HIV replication.