RESUMO
The chemokine macrophage inflammatory protein-1 alpha [MIP-1alpha] causes migration of B cells and also induces changes in antibody secretion. However, the signal transduction pathways leading to these phenotypic changes remain undefined. We have identified a signal transduction pathway initiated by MIP-1alpha in B cells. Here we report that stimulation of tonsil B cells with MIP-1alpha induces phosphatidylinositol 3-kinase [PI3-K] activation. Kinase activity was transient with peak induction occurring within 2.5 to 5 min after stimulation and was dose-dependent. In addition stimulation with MIP-1alpha induces tyrosine phosphorylation of the proline-rich tyrosine kinase Pyk-2. Immunoprecipitation analysis showed a constitutive association between Pyk-2 and PI3-K and pretreatment of MIP-1alpha-stimulated B cells with wortmannin, a specific inhibitor of PI3-K, resulted in a loss of PI3-K activity. The PI3-K inhibitor wortmannin prevented B cells from migrating in response to MIP-1alpha. Hence, PI3-K and Pyk-2 seem to be components of a signal transduction pathway induced by stimulation of B cells with MIP-1alpha, and this pathway may play a role in B-cell migration.
Assuntos
Linfócitos B/metabolismo , Movimento Celular/fisiologia , Proteínas Inflamatórias de Macrófagos/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Tirosina Quinases/metabolismo , Androstadienos/farmacologia , Linfócitos B/citologia , Movimento Celular/efeitos dos fármacos , Células Cultivadas , Quimiocina CCL3 , Quimiocina CCL4 , Relação Dose-Resposta a Droga , Indução Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Quinase 2 de Adesão Focal , Humanos , Proteínas Inflamatórias de Macrófagos/farmacologia , Tonsila Palatina , Inibidores de Fosfoinositídeo-3 Quinase , Fosforilação/efeitos dos fármacos , Testes de Precipitina , Ligação Proteica/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , WortmaninaRESUMO
Stimulation with the combination of PDB plus ionomycin induced LFA-1/ICAM-1-dependent homotypic adhesion of tonsil B cells. Adhesion of tonsil B cells in our system induced tyrosine phosphorylation of Pyk-2. Disruption of homotypic adhesion and concomitant inhibition of induction of protein tyrosine phosphorylation was achieved by physical separation of the cells and by treatment with methyl-2.5-dihydroxycinnamate (MDHC), an inhibitor of protein tyrosine phosphorylation. These results suggest that protein tyrosine phosphorylation that is associated with homotypic adhesion is mediated by LFA-1/ICAM-1 interactions.
Assuntos
Linfócitos B/enzimologia , Molécula 1 de Adesão Intercelular/fisiologia , Antígeno-1 Associado à Função Linfocitária/fisiologia , Proteínas Tirosina Quinases/metabolismo , Linfócitos B/metabolismo , Adesão Celular/imunologia , Agregação Celular/imunologia , Células Cultivadas , Inibidores Enzimáticos/farmacologia , Quinase 2 de Adesão Focal , Humanos , Tonsila Palatina , Fosforilação/efeitos dos fármacos , Fosfotirosina/metabolismo , Proteínas Tirosina Quinases/antagonistas & inibidoresRESUMO
The chemokine macrophage inflammatory protein-1 alpha (MIP-1alpha) stimulates migration of B cells through an unknown mechanism. Furthermore, little is known about signal transduction mechanisms through which MIP-1alpha might signal phenotypic changes in B cells. We are investigating the role of MIP-1alpha in B cell signaling. Here we report that stimulation of the Ramos B cell line or tonsil B or peripheral blood-B (PBL-B) cells with MIP-1alpha caused the transcription factor NF-kappaB to bind to DNA. NF-kappaB induction was dose dependent, and was transient, with peak induction occurring at 30 min. MIP-1alpha treatment stimulated the degradation of IkappaBalpha, a cytoplasmic inhibitor of NF-kappaB. The biological significance of NF-kappaB activation by MIP-1alpha is currently unknown, but it is known that NF-kappaB modulates expression of genes involved in many inflammatory and immune responses. Here we show that NF-kappaB activation is a target of signals sent into B cells by MIP-1alpha.
