CCR1-Mediated STAT3 Tyrosine Phosphorylation and CXCL8 Expression in THP-1 Macrophage-like Cells Involve Pertussis Toxin-Insensitive Gα14/16 Signaling and IL-6 Release

Agonists of CCR1 contribute to hypersensitivity reactions and atherosclerotic lesions, possibly via the regulation of the transcription factor STAT3. CCR1 was demonstrated to use pertussis toxin-insensitive G alpha(14/16) to stimulate phospholipase C beta and NF-kappa B, whereas both G alpha(14) and G alpha(16) are also capable of activating STAT3. The coexpression of CCR1 and G alpha(14/16) in human THP-1 macrophage-like cells suggests that CCR1 may use G alpha(14/16) to induce STAT3 activation. In this study, we demonstrated that a CCR1 agonist, leukotactin-1 (CCL15), could indeed stimulate STAT3 Tyr(705) and Ser(727) phosphorylation via pertussis toxin-insensitive G proteins in PMA-differentiated THP-1 cells, human erythroleukemia cells, and HEK293 cells overexpressing CCR1 and G alpha(14/16). The STAT3 Tyr(705) and Ser(727) phosphorylations were independent of each other and temporally distinct. Subcellular fractionation and confocal microscopy illustrated that Tyr(705)-phosphorylated STAT3 translocated to the nucleus, whereas Ser(727)-phosphorylated STAT3 was retained in the cytosol after CCR1/G alpha(14) activation. CCL15 was capable of inducing IL-6 and IL-8 (CXCL8) production in both THP-1 macrophage-like cells and HEK293 cells overexpressing CCR1 and G alpha(14/16). Neutralizing Ab to IL-6 inhibited CCL15-mediated STAT3 Tyr(705) phosphorylation, whereas inhibition of STAT3 activity abolished CCL15-activated CXCL8 release. The ability of CCR1 to signal through G alpha(14/16) provides a linkage for CCL15 to regulate IL-6/STAT3-signaling cascades, leading to expression of CXCL8, a cytokine that is involved in inflammation and the rupture of atherosclerotic plaque. The Journal of Immunology, 2012, 189: 5266-5276.