Activation and regulation of platelet-activating factor receptor: Role of Gi and Gq in receptor-mediated chemotactic, cytotoxic, and cross-regulatory signals

Stephan L. Brown, Venkatakrishna R. Jala, Sandeep K. Raghuwanshi, Mohd W. Nasser, Bodduluri Haribabu, Ricardo M. Richardson

Research output: Contribution to journalArticle

35 Citations (Scopus)

Abstract

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 Gi and Gq in PAF-induced leukocyte responses, two G-protein-linked receptors were generated by fusing Gαi3 (PAFR-Gαi3) or Gαq (PAFR-Gαq) at the C terminus of PAFR. Rat basophilic leukemia cell line (RBL-2H3) stably expressing wild-type PAFR, PAFR-Gαi3, or PAFR-Gαq 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 β-hexosaminidase. PAFR-Gαi3 and PAFR-Gα4 mediated greater GTPase activity in isolated membranes than PAFR but lower PI hydrolysis and secretion in whole cells. PAFR and PAFR-Gαi3, but not PAFR-Gαq, mediated chemotaxis to PAF. All three receptors underwent phosphorylation and desensitization upon exposure to PAF but only PAFR translocated βarrestin 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-Gαi3 activation cross-inhibited the response of CXCR1 to CXCL8. Altogether, the data indicate that Gi exclusively mediates chemotactic and cross-regulatory signals of the PAFR, but both Gi and Gq activate PI hydrolysis and exocytosis by this receptor. Because chemotaxis and cross-desensitization are exclusively mediated by Gi, the data suggest that differential activation of both Gi and G q by PAFR likely mediate specific as well as redundant signaling pathways.

Original languageEnglish (US)
Pages (from-to)3242-3249
Number of pages8
JournalJournal of Immunology
Volume177
Issue number5
DOIs
StatePublished - Sep 1 2006

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Phosphatidylinositols
GTP-Binding Proteins
Hydrolysis
Leukocytes
Pertussis Toxin
Chemotaxis
Interleukin-8
Arrestin
Hexosaminidases
Platelet Activating Factor
GTP Phosphohydrolases
Exocytosis
Cell Surface Receptors
Leukemia
Phosphorylation
Cell Membrane
Inflammation
Cell Line
Membranes
platelet activating factor receptor

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

Cite this

Activation and regulation of platelet-activating factor receptor : Role of Gi and Gq in receptor-mediated chemotactic, cytotoxic, and cross-regulatory signals. / Brown, Stephan L.; Jala, Venkatakrishna R.; Raghuwanshi, Sandeep K.; Nasser, Mohd W.; Haribabu, Bodduluri; Richardson, Ricardo M.

In: Journal of Immunology, Vol. 177, No. 5, 01.09.2006, p. 3242-3249.

Research output: Contribution to journalArticle

Brown, Stephan L. ; Jala, Venkatakrishna R. ; Raghuwanshi, Sandeep K. ; Nasser, Mohd W. ; Haribabu, Bodduluri ; Richardson, Ricardo M. / Activation and regulation of platelet-activating factor receptor : Role of Gi and Gq in receptor-mediated chemotactic, cytotoxic, and cross-regulatory signals. In: Journal of Immunology. 2006 ; Vol. 177, No. 5. pp. 3242-3249.
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abstract = "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 Gi and Gq in PAF-induced leukocyte responses, two G-protein-linked receptors were generated by fusing Gαi3 (PAFR-Gαi3) or Gαq (PAFR-Gαq) at the C terminus of PAFR. Rat basophilic leukemia cell line (RBL-2H3) stably expressing wild-type PAFR, PAFR-Gαi3, or PAFR-Gαq 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 β-hexosaminidase. PAFR-Gαi3 and PAFR-Gα4 mediated greater GTPase activity in isolated membranes than PAFR but lower PI hydrolysis and secretion in whole cells. PAFR and PAFR-Gαi3, but not PAFR-Gαq, mediated chemotaxis to PAF. All three receptors underwent phosphorylation and desensitization upon exposure to PAF but only PAFR translocated βarrestin 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-Gαi3 activation cross-inhibited the response of CXCR1 to CXCL8. Altogether, the data indicate that Gi exclusively mediates chemotactic and cross-regulatory signals of the PAFR, but both Gi and Gq activate PI hydrolysis and exocytosis by this receptor. Because chemotaxis and cross-desensitization are exclusively mediated by Gi, the data suggest that differential activation of both Gi and G q by PAFR likely mediate specific as well as redundant signaling pathways.",
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AU - Raghuwanshi, Sandeep K.

AU - Nasser, Mohd W.

AU - Haribabu, Bodduluri

AU - Richardson, Ricardo M.

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