Activation and regulation of platelet-activating factor receptor: role of G(i) and G(q) in receptor-mediated chemotactic, cytotoxic, and cross-regulatory signals

• Published in: The Journal of immunology (1950) 2006-09, Vol.177 (5), p.3242-3249
• Main Authors: Brown, Stephan L, Jala, Venkatakrishna R, Raghuwanshi, Sandeep K, Nasser, Mohd W, Haribabu, Bodduluri, Richardson, Ricardo M
• Format: Article
• Language: English
• Subjects: Animals; Calcium - metabolism; Cell Line, Tumor; Chemotaxis; Enzyme Activation; GTP-Binding Protein alpha Subunits, Gi-Go - metabolism; GTP-Binding Protein alpha Subunits, Gq-G11 - metabolism; Mitogen-Activated Protein Kinase 1 - metabolism; Mitogen-Activated Protein Kinase 3 - metabolism; Pertussis Toxin - pharmacology; Phosphorylation - drug effects; Platelet Activating Factor - pharmacology; Platelet Membrane Glycoproteins - metabolism; Protein Binding; Rats; Receptors, G-Protein-Coupled - metabolism; Receptors, Interleukin-8A - metabolism; Signal Transduction - drug effects
• ISSN: 0022-1767
• DOI: 10.4049/jimmunol.177.5.3242

Platelet-activating factor (1-O-alkyl-2-acetyl-sn-glycerolphosphocholine; PAF) induces leukocyte accumulation and activation at sites of inflammation via the activation of a specific cell surface receptor (PAFR). PAFR couples to both pertussis toxin-sensitive and pertussis toxin-insensitive G proteins to activate leukocytes. To define the role(s) of G(i) and G(q) in PAF-induced leukocyte responses, two G-protein-linked receptors were generated by fusing G alpha(i3) (PAFR-G alpha(i3)) or G alpha(q) (PAFR-G alpha(q)) at the C terminus of PAFR. Rat basophilic leukemia cell line (RBL-2H3) stably expressing wild-type PAFR, PAFR-G alpha(i3), or PAFR-G alpha(q) was generated and characterized. All receptor variants bound PAF with similar affinities to mediate G-protein activation, intracellular Ca2+ mobilization, phosphoinositide (PI) hydrolysis, and secretion of beta-hexosaminidase. PAFR-G alpha(i3) and PAFR-G alpha(q) mediated greater GTPase activity in isolated membranes than PAFR but lower PI hydrolysis and secretion in whole cells. PAFR and PAFR-G alpha(i3), but not PAFR-G alpha(q), mediated chemotaxis to PAF. All three receptors underwent phosphorylation and desensitization upon exposure to PAF but only PAFR translocated beta arrestin to the cell membrane and internalized. In RBL-2H3 cells coexpressing the PAFRs along with CXCR1, IL-8 (CXCL8) cross-desensitized Ca2+ mobilization to PAF by all the receptors but only PAFR-G alpha(i3) activation cross-inhibited the response of CXCR1 to CXCL8. Altogether, the data indicate that G(i) exclusively mediates chemotactic and cross-regulatory signals of the PAFR, but both G(i) and G(q) activate PI hydrolysis and exocytosis by this receptor. Because chemotaxis and cross-desensitization are exclusively mediated by G(i), the data suggest that differential activation of both G(i) and G(q) by PAFR likely mediate specific as well as redundant signaling pathways.