TDAG8 Is a Proton-sensing and Psychosine-sensitive G-protein-coupled Receptor
T cell death-associated gene 8 (TDAG8) has been reported to be a receptor for psychosine. Ovarian cancer G-protein-coupled receptor 1 (OGR1) and GPR4, G-protein-coupled receptors (GPCRs) closely related to TDAG8, however, have recently been identified as proton-sensing or extracellular pH-responsive GPCRs that stimulate inositol phosphate and cAMP production, respectively. In the present study, we examined whether TDAG8 senses extracellular pH change. In the several cell types that were transfected with TDAG8 cDNA, cAMP was markedly accumulated in response to neutral to acidic extracellular pH, with a peak response at approximately pH 7.0–6.5. The pH effect was inhibited by copper ions and was reduced or lost in cells expressing mutated TDAG8 in which histidine residues were changed to phenylalanine. In the membrane fractions prepared from TDAG8-transfected cells, guanosine 5′-O-(3-thiotriphosphate) binding activity and adenylyl cyclase activity were remarkably stimulated in response to neutral and acidic pH. The concentration-dependent effect of extracellular protons on cAMP accumulation was shifted to the right in the presence of psychosine. The inhibitory psychosine effect was also observed for pH-dependent actions in OGR1- and GPR4-expressing cells but not for prostaglandin E2- and sphingosine 1-phosphate-induced actions in any pH in native and sphingosine 1-phosphate receptor-expressing cells. Glucosylsphingosine and sphingosylphosphorylcholine similarly inhibited the pH-dependent action, although to a lesser extent. Psychosine-sensitive and pH-dependent cAMP accumulation was also observed in mouse thymocytes. We concluded that TDAG8 is one of the proton-sensing GPCRs coupling to adenylyl cyclase and psychosine, and its related lysosphingolipids behave as if they were antagonists against protein-sensing receptors, including TDAG8, GPR4, and OGR1. T cell death-associated gene 8 (TDAG8) has been reported to be a receptor for psychosine. Ovarian cancer G-protein-coupled receptor 1 (OGR1) and GPR4, G-protein-coupled receptors (GPCRs) closely related to TDAG8, however, have recently been identified as proton-sensing or extracellular pH-responsive GPCRs that stimulate inositol phosphate and cAMP production, respectively. In the present study, we examined whether TDAG8 senses extracellular pH change. In the several cell types that were transfected with TDAG8 cDNA, cAMP was markedly accumulated in response to neutral to acidic extracellular pH, with a peak response at approximately pH 7.0–6.5. The pH effect was inhibited by copper ions and was reduced or lost in cells expressing mutated TDAG8 in which histidine residues were changed to phenylalanine. In the membrane fractions prepared from TDAG8-transfected cells, guanosine 5′-O-(3-thiotriphosphate) binding activity and adenylyl cyclase activity were remarkably stimulated in response to neutral and acidic pH. The concentration-dependent effect of extracellular protons on cAMP accumulation was shifted to the right in the presence of psychosine. The inhibitory psychosine effect was also observed for pH-dependent actions in OGR1- and GPR4-expressing cells but not for prostaglandin E2- and sphingosine 1-phosphate-induced actions in any pH in native and sphingosine 1-phosphate receptor-expressing cells. Glucosylsphingosine and sphingosylphosphorylcholine similarly inhibited the pH-dependent action, although to a lesser extent. Psychosine-sensitive and pH-dependent cAMP accumulation was also observed in mouse thymocytes. We concluded that TDAG8 is one of the proton-sensing GPCRs coupling to adenylyl cyclase and psychosine, and its related lysosphingolipids behave as if they were antagonists against protein-sensing receptors, including TDAG8, GPR4, and OGR1. TDAG8 1The abbreviations used are: TDAG8, T cell death-associated gene 8; GPCR, G-protein-coupled receptor; OGR1, ovarian cancer G-protein-coupled receptor 1; SPC, sphingosylphosphorylcholine; LPC, l-α-lysophosphatidylcholine; PGE2, prostaglandin E2; S1P, sphingosine 1-phosphate; BSA, bovine serum albumin; RT, reverse transcription; CHO, Chinese hamster ovary; WT, wild type; DMEM, Dulbecco's modified Eagle's medium; PTX, pertussis toxin; DEX, dexamethasone; IBMX, 3-isobutyl-1-methylxanthine; EPPS, N′-(2-hydroxyethyl)piperazine-N′-3-propanesulfonic acid; MES, 2-(N-morpholine)ethanesulfonic acid; GTPγS, guanosine 5′-O-(3-thiotriphosphate); S1P3, S1P3/EDG-3 receptor.1The abbreviations used are: TDAG8, T cell death-associated gene 8; GPCR, G-protein-coupled receptor; OGR1, ovarian cancer G-protein-coupled receptor 1; SPC, sphingosylphosphorylcholine; LPC, l-α-lysophosphatidylcholine; PGE2, prostaglandin E2; S1P, sphingosine 1-phosphate; BSA, bovine serum albumin; RT, reverse transcription; CHO, Chinese hamster ovary; WT, wild type; DMEM, Dulbecco's modified Eagle's medium; PTX, pertussis toxin; DEX, dexamethasone; IBMX, 3-isobutyl-1-methylxanthine; EPPS, N′-(2-hydroxyethyl)piperazine-N′-3-propanesulfonic acid; MES, 2-(N-morpholine)ethanesulfonic acid; GTPγS, guanosine 5′-O-(3-thiotriphosphate); S1P3, S1P3/EDG-3 receptor. was initially cloned as an orphan GPCR, which is up-regulated during the programmed cell death of T lymphocytes (1Choi J.W. Lee S.Y. Choi Y. Cell. Immunol. 1996; 168: 78-84Crossref PubMed Scopus (87) Google Scholar, 2Kyaw H. Zeng Z. Su K. Fan P. Shell B.K. Carter K.C. Li Y. DNA Cell Biol. 1998; 17: 493-500Crossref PubMed Scopus (47) Google Scholar, 3Tosa N. Murakami M. Jia W.Y. Yokoyama M. Masunaga T. Iwabuchi C. Inobe M. Iwabuchi K. Miyazaki T. Onoe K. Iwata M. Uede T. Int. Immunol. 2003; 15: 741-749Crossref PubMed Scopus (51) Google Scholar). This gene product has recently been reported (4Im D.S. Heise C.E. Nguyen T. O'Dowd B.F. Lynch K.R. J. Cell Biol. 2001; 153: 429-434Crossref PubMed Scopus (152) Google Scholar) to be a receptor for psychosine, a lysosphingolipid, which induces the formation of multinuclear cells. OGR1, which shares 41% identical amino acids with TDAG8, was initially reported (5Xu Y. Zhu K. Hong G. Wu W. Baudhuin L.M. Xiao Y. Damron D.S. Nat. Cell Biol. 2000; 2: 261-267Crossref PubMed Scopus (173) Google Scholar) to be a receptor for sphingosylphosphorylcholine (SPC). GPR4 also shares homology with TDAG8 and was identified as a receptor for lysolipids, including lysophosphatidylcholine (LPC) and SPC (6Zhu K. Baudhuin L.M. Hong G. Williams F.S. Cristina K.L. Kabarowski J.H. Witte O.N. Xu Y. J. Biol. Chem. 2001; 276: 41325-41335Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar). It has recently been reported (7Ludwig M.G. Vanek M. Guerini D. Gasser J.A. Jones C.E. Junker U. Hofstetter H. Wolf R.M. Seuwen K. Nature. 2003; 425: 93-98Crossref PubMed Scopus (501) Google Scholar), however, that OGR1 and GPR4 sense extracellular protons through histidine residues of receptors and are coupled to G-proteins to stimulate intracellular signaling pathways. Thus, OGR1 stimulation causes inositol phosphate production, and the subsequent mobilization of intracellular calcium and GPR4 stimulation induces cAMP accumulation, probably reflecting the activation of adenylyl cyclase in response to an extracellular pH change (7Ludwig M.G. Vanek M. Guerini D. Gasser J.A. Jones C.E. Junker U. Hofstetter H. Wolf R.M. Seuwen K. Nature. 2003; 425: 93-98Crossref PubMed Scopus (501) Google Scholar). These results raise the possibility that TDAG8 may also respond to extracellular pH change and stimulate intracellular signaling pathways. If TDAG8 is proven to be one of the pH-sensing receptors, the next issue is the relationship between two putative ligands, lysolipid and proton, with respect to the action mechanism; that is, do the two ligands recognize these proton-sensing receptors independently of each other? In a previous study (7Ludwig M.G. Vanek M. Guerini D. Gasser J.A. Jones C.E. Junker U. Hofstetter H. Wolf R.M. Seuwen K. Nature. 2003; 425: 93-98Crossref PubMed Scopus (501) Google Scholar), the specific action of SPC, a putative OGR1 agonist, could not be observed in OGR1-expressing cells. In the present study, we observed that TDAG8 senses extracellular pH and stimulates adenylyl cyclase and the subsequent accumulation of cellular cAMP. Mutagenesis experiments on TDAG8 suggest that histidine residues may play a role in proton sensing. Furthermore, we observed that psychosine and its related lysosphingolipids inhibited the proton-sensing responses in a manner that is dependent on extracellular pH in the cells expressing not only TDAG8 but also OGR1 and GPR4. Thus, TDAG8 is one of the proton-sensing GPCRs that is sensitive to psychosine and its related lysosphingolipids. These lipids behave as if they were antagonists for the proton-sensing GPCRs and may be useful tools for characterization of this group of receptors. Materials—Psychosine (galactosylsphingosine), glucosylsphingosine, SPC, d-erythro-sphingosine, and prostaglandin E2 (PGE2) were purchased from Sigma. 1-Oleoyl-sn-glycero-3-phosphate (lysophosphatidic acid), l-α-lysophosphatidylcholine palmitoyl (LPC, C16:0), and sphingosine 1-phosphate (S1P) were from Cayman Chemical Co. (Ann Arbor, MI). Fatty acid-free BSA was from Calbiochem-Novabiochem; PTX was from List Biological Laboratories, Inc. (Campbell, CA); anti-Gαs/Gαolf (C-18) antibody, a polyclonal antibody raised against a peptide mapping at the C-terminal Gαs, and the C-terminal peptide for the blocking were from Santa Cruz Biotechnology (Santa Cruz, CA); and myo-[2-3H]inositol (23.0 Ci/mmol) was from American Radiolabeled Chemicals, Inc. (St. Louis, MO). A 14-peptide fragment of muscarinic receptor m4I3C was synthesized as described previously (8Kuribayashi F. Nunoi H. Wakamatsu K. Tsunawaki S. Sato K. Ito T. Sumimoto H. EMBO J. 2002; 21: 6312-6320Crossref PubMed Scopus (122) Google Scholar). Mice—C57BL/6 mice (females, 5 weeks of age) were obtained from Charles River Japan, Inc. (Tokyo, and were to the mice The mice were 1 to by the of and of TDAG8 OGR1 and GPR4 were from a by The fragment was the of a J. Sato K. T. H. T. K. T. N. T. H. M. F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, K. M. F. 2000; PubMed Scopus Google Scholar) for TDAG8 and OGR1, and a for GPR4. DNA was The an a and the of the receptor. The an and a in to the C-terminal of the receptor. cells or cells were transfected with the by and the at 1 for cells and for cells were cells to were used in cells expressing TDAG8 receptor were cloned by the two cells expressing TDAG8 or were were by and cloned of to or in which the or of histidine from the was changed to respectively. TDAG8 was also cloned to These mutated or wild TDAG8 were transfected cells by to in the cells H. H. K. Sato K. M. M. Y. F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Cell cell of cells and which were transfected with and or the receptor J. Sato K. T. H. T. K. T. N. T. H. M. F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar), and were in bovine serum in a cAMP and inositol phosphate the cells were on the the was changed to BSA for the cAMP response and to of 1 and PTX of the cells was by the to the the cells transfected with and were for in bovine and the was with BSA the were in of with bovine serum in the presence or of 1 the and cells were and used for the of and cAMP of cAMP in cells, for were and for at in of 5 and BSA The cells were for the pH in the presence of in a of a pH the was with a were with the in this are to the pH at The was by of 1 the was used for the cAMP response in CHO, and cells, we not any between the response of the and the at in the cAMP response in cells. In the of the cells were with the and with the The cell were to the which IBMX, the pH and were for at The was by in the was as described previously J. Sato K. T. H. T. K. T. N. T. H. M. F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). cells transfected with the receptors were and with the for The cells were for at in the pH in the presence of inositol including inositol and were as described previously J. Sato K. T. H. T. K. T. N. T. H. M. F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). membrane fractions were prepared from and TDAG8-transfected cells with and as described previously J. Sato K. T. H. T. K. T. N. T. H. M. F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The binding was by the with for at as described previously H. Sato K. N. J. T. M. Y. T. M. Sato M. H. T. T. D.S. H. K. H. F. 2003; PubMed Scopus Google Scholar), that the pH of the was In of at pH were with the the pH in of 5 and The was by of an by an through a The was with the and at for 1 for in a membrane fractions that were prepared for the binding were used for the adenylyl cyclase The in were with the the pH in of IBMX, GTPγS, and in 5 and for at In the in which the effect of antibody for was were with the antibody for at was by of 1 was to the to of DNA the 5 of the was and reverse to the the of the OGR1 TDAG8 and GPR4 was with a The mouse and were obtained from for OGR1, for TDAG8, and for sense and of mouse were synthesized as and respectively. were described previously T. Sato K. M. M. T. Y. T. Y. T. S. Y. H. D.S. K. H. F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The of the was to the of the of was at and the results are as the experiments were in or The results of are as the or as results from of cells was by the were at TDAG8 pH in cAMP examined cAMP accumulation in cells transfected with and TDAG8 adenylyl cyclase IBMX, an of was in the in in cells a acidic pH of not change the cAMP and and cAMP accumulation with accumulation at pH Thus, of extracellular pH to the activity of or the activity of adenylyl the in the TDAG8-transfected cells, the acidic pH remarkably the cAMP and were to the The response was observed at pH The cAMP response to pH was to PTX of which the of cAMP accumulation in the presence of not These results suggest that are not in the proton-sensing the extracellular pH change a inositol phosphate accumulation in cells, but effect on inositol phosphate response was by a pH change in or TDAG8-transfected cells inhibited pH-dependent responses in the cells transfected with proton-sensing and TDAG8-transfected cells were for inositol phosphate and cAMP responses to the pH in the presence or of psychosine. TDAG8 cells were at pH in the presence of the of psychosine SPC, LPC, and sphingosine cells were the pH in the presence or of psychosine. GPR4 we used of however, cells not respond to extracellular pH not cells were for inositol phosphate response to pH in the presence or of The effect of the extracellular pH change in cAMP accumulation was also observed in cells transfected with TDAG8 cDNA, effect was in response to the pH change in cells if the cAMP response the activity of adenylyl is that TDAG8 receptor stimulation the through intracellular and extracellular signaling this we prepared from these cells and binding activity and adenylyl cyclase activity in the in a change in pH in the stimulated binding in the probably reflecting the in the binding to Thus, a proton as a receptor for In these membrane from cloned cells, the activity of the of the adenylyl cyclase to be identical to that of the cell as from the that the activity by the or a 14-peptide fragment of muscarinic receptor (8Kuribayashi F. Nunoi H. Wakamatsu K. Tsunawaki S. Sato K. Ito T. Sumimoto H. EMBO J. 2002; 21: 6312-6320Crossref PubMed Scopus (122) Google Scholar), was at pH in proton in the however, a in the activity to pH and the activity with the of the which was probably by the of the of the for the with proton the activity was reduced in prepared from the cells may the adenylyl cyclase activation as by a of the activation by anti-Gαs/Gαolf antibody The inhibitory of the antibody were by with the peptide not This antibody has been to recognize but not or D. J. PubMed Scopus Google Scholar). Thus, TDAG8 receptors to to G-proteins and probably and a proton may the and the subsequent activation of adenylyl In the of OGR1-expressing cells, was in the probably by the of histidine (7Ludwig M.G. Vanek M. Guerini D. Gasser J.A. Jones C.E. Junker U. Hofstetter H. Wolf R.M. Seuwen K. Nature. 2003; 425: 93-98Crossref PubMed Scopus (501) Google Scholar). The proton-sensing cAMP response in cells was inhibited by the response was by the The of histidine residues was also by experiments in which cAMP accumulation in cells transfected with the wild or mutated of TDAG8 was in a at a of the histidine residues or in TDAG8 inhibited the cAMP accumulation with a activity and a The change in the not to be the of the of the receptor a in the of transfected wild DNA changed the activity but not change the The in is to in OGR1 as is to in GPR4 (7Ludwig M.G. Vanek M. Guerini D. Gasser J.A. Jones C.E. Junker U. Hofstetter H. Wolf R.M. Seuwen K. Nature. 2003; 425: 93-98Crossref PubMed Scopus (501) Google Scholar), and are in extracellular Thus, these extracellular histidine residues may play a role in extracellular the is to in OGR1 and is in the The in this in a in the proton-sensing response as was the in OGR1, that the may be for (7Ludwig M.G. Vanek M. Guerini D. Gasser J.A. Jones C.E. Junker U. Hofstetter H. Wolf R.M. Seuwen K. Nature. 2003; 425: 93-98Crossref PubMed Scopus (501) Google of on TDAG8 receptors in of proton-sensing cAMP response in cells. cells were transfected with of or and cAMP response to the pH was the cells were transfected with or of the was or of that observed in transfected cells the the was pH and for cells transfected with and of by and of has recently been reported (4Im D.S. Heise C.E. Nguyen T. O'Dowd B.F. Lynch K.R. J. Cell Biol. 2001; 153: 429-434Crossref PubMed Scopus (152) Google Scholar) to be a receptor for psychosine. OGR1 and GPR4 have also been to be receptors for as SPC and (5Xu Y. Zhu K. Hong G. Wu W. Baudhuin L.M. Xiao Y. Damron D.S. Nat. Cell Biol. 2000; 2: 261-267Crossref PubMed Scopus (173) Google Scholar, K. Baudhuin L.M. Hong G. Williams F.S. Cristina K.L. Kabarowski J.H. Witte O.N. Xu Y. J. Biol. Chem. 2001; 276: 41325-41335Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar). in however, we to a effect of psychosine on inositol phosphate response at any pH in TDAG8-transfected cells. the the concentration-dependent of the proton was shifted to the right in the presence of psychosine Thus, psychosine as if was an TDAG8 was stimulated with the lysolipids, including and SPC, although to a lesser psychosine, also inhibited the cAMP response to the neutral pH in TDAG8-transfected cells. the of and sphingosine were or at In the previous the of psychosine on OGR1 and GPR4 have not been however, the of the psychosine effect was also observed for cAMP accumulation in cells and inositol phosphate accumulation in cells although psychosine to be for GPR4 with TDAG8 and OGR1. The inhibitory effect of psychosine was specific to the proton-sensing receptors. Thus, cAMP accumulation through expressing receptors was by psychosine at pH of the of TDAG8 A and was with psychosine was not not at pH psychosine was for the cAMP response through and receptors. The cAMP response to in cells was also to psychosine at pH not GPCRs cAMP in is in including the and (1Choi J.W. Lee S.Y. Choi Y. Cell. Immunol. 1996; 168: 78-84Crossref PubMed Scopus (87) Google Scholar, 2Kyaw H. Zeng Z. Su K. Fan P. Shell B.K. Carter K.C. Li Y. DNA Cell Biol. 1998; 17: 493-500Crossref PubMed Scopus (47) Google Scholar, 3Tosa N. Murakami M. Jia W.Y. Yokoyama M. Masunaga T. Iwabuchi C. Inobe M. Iwabuchi K. Miyazaki T. Onoe K. Iwata M. Uede T. Int. Immunol. 2003; 15: 741-749Crossref PubMed Scopus (51) Google Scholar). In the has been to the of TDAG8 H. Zeng Z. Su K. Fan P. Shell B.K. Carter K.C. Li Y. DNA Cell Biol. 1998; 17: 493-500Crossref PubMed Scopus (47) Google Scholar, 3Tosa N. Murakami M. Jia W.Y. Yokoyama M. Masunaga T. Iwabuchi C. Inobe M. Iwabuchi K. Miyazaki T. Onoe K. Iwata M. Uede T. Int. Immunol. 2003; 15: 741-749Crossref PubMed Scopus (51) Google Scholar). We examined the of proton-sensing and cAMP response to extracellular pH in mouse thymocytes. with the previous results H. Zeng Z. Su K. Fan P. Shell B.K. Carter K.C. Li Y. DNA Cell Biol. 1998; 17: 493-500Crossref PubMed Scopus (47) Google Scholar, 3Tosa N. Murakami M. Jia W.Y. Yokoyama M. Masunaga T. Iwabuchi C. Inobe M. Iwabuchi K. Miyazaki T. Onoe K. Iwata M. Uede T. Int. Immunol. 2003; 15: 741-749Crossref PubMed Scopus (51) Google Scholar), markedly the of TDAG8 in which was observed at and the We also the of of OGR1 and GPR4 A GPR4 was also at the and its was in response to although a effect was not the and the of the effect was that of for OGR1, a of was not at the and the effect was The effect of extracellular pH on cAMP response was examined in to acidic extracellular pH, in a in cAMP In with the in the of proton-sensing receptor TDAG8 and GPR4, by the cAMP response to acidic pH was by the It is that the of the cAMP response was at the was a in of TDAG8 but not in GPR4 by the the and cAMP responses were inhibited by The effect of psychosine on cAMP response is in and The in cAMP accumulation was inhibited by psychosine the cAMP response to or was by psychosine These results suggest that an acidic in cAMP accumulation may be by proton-sensing GPCRs and that TDAG8, may play an In the present study, we have that TDAG8 is a proton-sensing and its stimulation results in cellular cAMP We have that psychosine and its related including SPC and glucosylsphingosine, cAMP accumulation as if these lipids were antagonists for the TDAG8 receptor the receptor is stimulated with was also for proton-sensing receptors, including OGR1 and GPR4, by the of histidine residues for activation of these receptors. on a study on OGR1, (7Ludwig M.G. Vanek M. Guerini D. Gasser J.A. Jones C.E. Junker U. Hofstetter H. Wolf R.M. Seuwen K. Nature. 2003; 425: 93-98Crossref PubMed Scopus (501) Google Scholar) that between histidine residues the of the receptors at a pH and that the of by at a acidic pH induces the change in the of the receptor in of coupling to may the acidic (7Ludwig M.G. Vanek M. Guerini D. Gasser J.A. Jones C.E. Junker U. Hofstetter H. Wolf R.M. Seuwen K. Nature. 2003; 425: 93-98Crossref PubMed Scopus (501) Google Scholar). The results of the present study on TDAG8 also suggest that histidine residues are in the proton-sensing although but not to to The of the concentration-dependent cAMP accumulation of an extracellular proton was approximately that a pH change induces a change in the receptor of TDAG8 was initially reported (4Im D.S. Heise C.E. Nguyen T. O'Dowd B.F. Lynch K.R. J. Cell Biol. 2001; 153: 429-434Crossref PubMed Scopus (152) Google Scholar) to be a receptor for psychosine, on the results that psychosine an of cAMP accumulation and an in intracellular we to an of cAMP accumulation by psychosine extracellular pH was Furthermore, psychosine was for inositol phosphate and for intracellular not we observed the inhibitory action of psychosine on cAMP the extracellular pH was neutral or acidic the for the between previous (4Im D.S. Heise C.E. Nguyen T. O'Dowd B.F. Lynch K.R. J. Cell Biol. 2001; 153: 429-434Crossref PubMed Scopus (152) Google Scholar) and present results be that the inhibitory action of psychosine on cAMP accumulation was at pH and of pH of psychosine action on the cAMP In any results are not in with the of psychosine as a for the TDAG8 receptor. to behave at as an of in cAMP. Glucosylsphingosine and SPC were also for cAMP accumulation if extracellular pH was neutral or acidic The action of psychosine on the pH-dependent response was not to the TDAG8 receptor but was to proton-sensing receptors, including OGR1 and GPR4 the present study is the to that psychosine may with OGR1 and GPR4. In the presence of psychosine, the concentration-dependent of the extracellular proton on cAMP accumulation or inositol phosphate shifted to the right We that psychosine, to with histidine residues and the or with the of histidine residues on receptor for the change to the and lysosphingolipids were examined for to binding and adenylyl cyclase activity in TDAG8-transfected cell we to a specific inhibitory action by these in the of the actions not These results also suggest that psychosine and on the cells from the are to the action of psychosine and related lysosphingolipids to the activation of proton-sensing In the previous study (4Im D.S. Heise C.E. Nguyen T. O'Dowd B.F. Lynch K.R. J. Cell Biol. 2001; 153: 429-434Crossref PubMed Scopus (152) Google Scholar) psychosine was to multinuclear formation in the cells expressing TDAG8 cells. It has not been however, whether of cells by psychosine is also by its action against the proton-sensing TDAG8 receptor. of TDAG8 as by has that the gene is in including the and (1Choi J.W. Lee S.Y. Choi Y. Cell. Immunol. 1996; 168: 78-84Crossref PubMed Scopus (87) Google Scholar, 2Kyaw H. Zeng Z. Su K. Fan P. Shell B.K. Carter K.C. Li Y. DNA Cell Biol. 1998; 17: 493-500Crossref PubMed Scopus (47) Google Scholar, 3Tosa N. Murakami M. Jia W.Y. Yokoyama M. Masunaga T. Iwabuchi C. Inobe M. Iwabuchi K. Miyazaki T. Onoe K. Iwata M. Uede T. Int. Immunol. 2003; 15: 741-749Crossref PubMed Scopus (51) Google Scholar). The of that TDAG8 may play an role in the In the present study, we have that acidic pH causes cAMP accumulation and that cAMP accumulation is inhibited by psychosine in mouse Furthermore, the cAMP response was at the which was to the of the of TDAG8 GPR4 was also by but the effect was not These results suggest that and proton-sensing GPCRs are in the of the cAMP response to acidic extracellular TDAG8 may be for this and GPR4 also play a role but in the the receptor however, is to the of each receptor. Furthermore, we the of an pH-dependent proton-sensing Thus, of the of TDAG8 and proton-sensing GPCRs tools to each receptor and its from or an for proton-sensing receptors may to these which is the of the role of proton-sensing receptors to be cAMP has been reported S. P. M. 2000; PubMed Google Scholar, J.A. J. J. Immunol. 2000; PubMed Scopus Google Scholar) to in thymocytes. Thus, proton-sensing GPCRs may play a role in the of to during in the Immunol. 2000; PubMed Scopus Google Scholar). The of TDAG8 is not to is also in and Thus, of the role of proton-sensing GPCRs in and not only a the of of or but also a for in and the of the present study, a with a to TDAG8, OGR1, and GPR4, was also reported to sense extracellular proton, to the of inositol phosphate and activation of signaling N. T. T. T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Thus, these receptors to a of proton-sensing We for