Tyrosine phosphorylation-mediated signal transduction in MCP-1-induced macrophage activation: role for receptor dimerization, focal adhesion protein complex and JAK/STAT pathway

Abstract

Monocyte chemoattractant protein-1 (MCP-1) plays a crucial role in the recruitment of monocytes/macrophages associated with several inflammatory diseases and malignancies. The early signal transduction mechanism of macrophage activation in response to in vitro MCP-1 treatment was investigated. The treatment of murine peritoneal macrophages with MCP-1 resulted in a significant enhancement in the tyrosine phosphorylation of cellular proteins, which peaked within 2.5–5 min of MCP-1 treatment. The MCP-1-induced tyrosine phosphorylation of cellular proteins involved the phosphorylation of non-receptor tyrosine kinases Lyn, JAK2, cytoskeletal binding protein paxillin and downstream transcription factors STAT3 and STAT5. Immunoflourescence microscopical studies on MCP-1-treated macrophages showed the cellular localization of the tyrosine-phosphorylated proteins and bundling of actin filaments at the focal adhesion points. MCP-1-induced association of focal adhesion proteins Lyn/phospho-paxillin with CCR2 was also observed by co-precipitation. Inhibitor studies with genistein on MCP-1-induced macrophage TNF and IL-1 production additionally supported the role of protein tyrosine phosphorylation in the process of macrophage activation with MCP-1. Present investigations suggest that the early events in the tyrosine kinase signal transduction pathway for macrophage activation in response to MCP-1 probably involve (1) CCR2 receptor dimerization, (2) enhanced tyrosine phosphorylation and assembly of focal adhesion complex, and (3) the activation of JAK/STAT pathway in the murine peritoneal macrophages.