Differential Regulation of CXCL8 Production by Different G Protein Subunits with Synergistic Stimulation by G_i- and G_q-Regulated Pathways

CXCL8 (also known as interleukin-8 or IL-8) is a proinflammatory chemokine that not only modulates the inflammatory and immune responses, but whose upregulation is often associated with diseases including various types of cancer. Although numerous ligands for G protein-coupled receptors (GPCRs) have been shown to stimulate the production of CXCL8, the specificity of the G protein signal remains undefined. By expressing the constitutively active Gα subunits in HEK293 cells, CXCL8 production was herein demonstrated to be most effectively stimulated by Gα_q family members, while those of Gα_s and Gα₁₂ elicited much weaker activities, and Gα_i being totally ineffective. However, in cell lines such as HepG2, HeLa, and MCF-7 that endogenously express Gβγ-responsive phospholipase Cβ isoforms (PLCβ_2/3), activation of the G_i-coupled α₂-adrenoceptor significantly stimulated CXCL8 production. This G_i-induced CXCL8 production was apparently mediated via specific Gβγ dimers and required the presence of PLCβ_2/3. Co-activation of G_i-coupled α₂-adrenoceptor and G_q-coupled bradykinin receptor resulted in a synergistic CXCL8 production, with Gβγ-responsive PLCβ_2/3, Src, ERK, and STAT3 serving as critical signaling intermediates. The treatment of HepG2 and B-10 endothelial cells with bradykinin stimulated CXCL8 production and cell proliferation. Interestingly, the latter response was driven by CXCL8 autocrine signaling because it was abolished by SB225002, an antagonist that prevents CXCL8 from binding to CXCR2. Collectively, our results provide a mechanistic basis for various G protein subfamilies to regulate the production of CXCL8, which may then lead to paracrine and/or autocrine signaling with major implications in both normal physiology and pathophysiological conditions.