Chemokines have been shown to induce and direct adhesion, chemotaxis, activation, and degranulation of human and rodent leukocytes both in vitro and in vivo. CXCL12 and CCL19 are two important chemokines that regulate T cell motility and activation under normal and inflammatory conditions. Despite numerous reports examining the function of chemokines, little is known about the molecular changes induced by these ligands. We have identified by microarray analysis differentially expressed genes in human T cells following treatment with CXCL12 or CCL19. In primary T cells, it was observed that the genes associated with non canonical Wnt signaling pathway are significantly induced by chemokines compared to vehicle control-treated cells. The non-canonical Wnts, Wnt5a and Wnt10a, along with their receptors Fzd2 and Fzd3 were significantly induced upon CXCL12 and CCL19 treatment, respectively, in a time-dependent fashion. The up regulation of these genes was confirmed by real time PCR and western blot analysis. In addition, the transcriptional activation of both Wnt5a and Wnt10a promoter by CXCL12 and CCL19 was further confirmed by electrophoretic mobility shift assay (EMSA). siRNA and shRNA knockdown of Wnt5a in primary T cells and transformed T cell lines, respectively, significantly inhibited CXCR4 and CCR7 signaling and function suggesting an important intermediary role for the Wnt-Fzd pathways in chemokine-chemokine receptor function. Moreover, these chemokine-induced Wnt and Fzd molecules appear to play a critical role in PKC and Rac activation, MMP expression and actin polymerization in response to chemokine stimulation. Similar effects were observed using melanoma cell lines. In vivo studies are currently underway to support the function role of this pathway in lymphocyte migration and trafficking. Together, these results provide insight into chemokine-induced gene activation and implicate the possible involvement of the non-canonical Wnt signaling pathway in chemokine activation and T cell trafficking. ? ? Moreover, we are also currently verifying and characterizing several additional gene families that are highly expressed in T cells after migration in response to or simply stimulation with CXCL12, CCL19, gp120 and HIV-1 virus. Moreover, the role of lipid rafts in chemokine biology and HIV infectivity are also under examination using microarray analysis. A greater understanding of the transcriptional signals differentially induced by the ligation of various chemokine receptors may provide a means to dissect the pathways by which these chemoattractants induce cell migration and activation as well as any host transcriptional signals important in HIV entry and replication.
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