title: Sphingosylphosphorylcholine and lysophosphatidylcholine are ligands for the G protein-coupled receptor GPR4.
abstract: Sphingosylphosphorylcholine (SPC) and lysophosphatidylcholine (LPC) are bioactive lipid molecules involved in numerous biological processes. We have recently identified ovarian cancer G protein-coupled receptor 1 (OGR1) as a specific and high affinity receptor for SPC, and G2A as a receptor with high affinity for LPC, but low affinity for SPC. Among G protein-coupled receptors, GPR4 shares highest sequence homology with OGR1 (51%). In this work, we have identified GPR4 as not only another high affinity receptor for SPC, but also a receptor for LPC, albeit of lower affinity. Both SPC and LPC induce increases in intracellular calcium concentration in GPR4-, but not vector-transfected MCF10A cells. These effects are insensitive to treatment with BN52021, WEB-2170, and WEB-2086 (specific platelet activating factor (PAF) receptor antagonists), suggesting that they are not mediated through an endogenous PAF receptor. SPC and LPC bind to GPR4 in GPR4-transfected CHO cells with K(d)/SPC = 36 nm, and K(d)/LPC = 159 nm, respectively. Competitive binding is elicited only by SPC and LPC. Both SPC and LPC activate GPR4-dependent activation of serum response element reporter and receptor internalization. Swiss 3T3 cells expressing GPR4 respond to both SPC and LPC, but not sphingosine 1-phosphate (S1P), PAF, psychosine (Psy), glucosyl-beta1'1-sphingosine (Glu-Sph), galactosyl-beta1'1-ceramide (Gal-Cer), or lactosyl-beta1'1-ceramide (Lac-Cer) to activate extracellular signal-regulated kinase mitogen-activated protein kinase in a concentration- and time-dependent manner. SPC and LPC stimulate DNA synthesis in GPR4-expressing Swiss 3T3 cells. Both extracellular signal-regulated kinase activation and DNA synthesis stimulated by SPC and LPC are pertussis toxin-sensitive, suggesting the involvement of a G(i)-heterotrimeric G protein. In addition, GPR4 expression confers chemotactic responses to both SPC and LPC in Swiss 3T3 cells. Taken together, our data indicate that GPR4 is a receptor with high affinity to SPC and low affinity to LPC, and that multiple cellular functions can be transduced via this receptor.
authors.affiliation: Department of Cancer Biology, Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
authors.forename: K
authors.initials: K
authors.lastname: Zhu
authors.forename: L M
authors.initials: LM
authors.lastname: Baudhuin
authors.forename: G
authors.initials: G
authors.lastname: Hong
authors.forename: F S
authors.initials: FS
authors.lastname: Williams
authors.forename: K L
authors.initials: KL
authors.lastname: Cristina
authors.forename: J H
authors.initials: JH
authors.lastname: Kabarowski
authors.forename: O N
authors.initials: ON
authors.lastname: Witte
authors.forename: Y
authors.initials: Y
authors.lastname: Xu
mesh_terms: D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D006224:Cricetinae; D017931:DNA Primers; D004261:DNA Replication; D004789:Enzyme Activation; D006801:Humans; D008024:Ligands; D008244:Lysophosphatidylcholines / Q000378:metabolism*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010767:Phosphorylcholine / Q000031:analogs & derivatives* / Q000378:metabolism*; D011869:Radioligand Assay; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000378:metabolism*; D014162:Transfection
chemical_list: D017931:DNA Primers; C091670:GPR4 protein, human; D008024:Ligands; D008244:Lysophosphatidylcholines; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; C005356:sphingosine phosphorylcholine; D010767:Phosphorylcholine; D020928:Mitogen-Activated Protein Kinases; D013110:Sphingosine; D002118:Calcium