Different signaling pathway between sphingosine-1-phosphate and lysophosphatidic acid in Xenopus oocytes: functional coupling of the sphingosine-1-phosphate receptor to PLC-xbeta in Xenopus oocytes.
In Xenopus oocytes, both sphingosine-1-phosphate (S1P) and lysophosphatidic acid (LPA) activate Ca2+-dependent oscillatory Cl- currents by acting through membrane-bound receptors. External application of 50 microM S1P elicited a long-lasting oscillatory current that continued over 30 min from the beginning of oscillation, with 300 nA (n = 11) as a usual maximum peak of current, whereas 1-microM LPA treatment showed only transiently oscillating but more vigorous current responses, with 2,800 nA (n = 18) as a maximum peak amplitude. Both phospholipid-induced Ca2+-dependent Cl- currents were observed in the absence of extracellular Ca2+, were blocked by intracellular injection of the Ca2+ chelator, EGTA, and could not be elicited by treatment with thapsigargin, an inhibitor of endoplasmic reticulum (ER) Ca2+ ATPase. Intracellular Ca2+ release appeared to be from inositol 1,4,5-trisphosphate (IP3)-sensitive Ca2+ store, because Cl- currents were blocked by heparin injection. Pretreatment with the aminosteroid, U-73122, an inhibitor of G protein-mediated phospholipase C (PLC) activation, to oocytes inhibited the current responses evoked both by S1P and LPA. However, when they were injected with 10 ng of antisense oligonucleotide (AS-ODN) against Xenopus phospholipase C (PLC-xbeta), oocytes could not respond to S1P application, whereas they responded normally to LPA, indicating that the S1P signaling pathway goes through PLC-xbeta, whereas LPA signaling goes through another unknown PLC. To determine the types of G proteins involved, we introduced AS-ODNs against four types of G-protein alpha subunits that were identified in Xenopus laevis; G(q)alpha, G11alpha, G0alpha, and G(i1)alpha. Among AS-ODNs against the G alphas tested, AS-G(q)alpha and AS-G(i1)alpha to S1P and AS-G(q)alpha and AS-G11alpha to LPA specifically reduced current responses, respectively, to about 20-30% of controls. These results demonstrate that LPA and S1P, although they have similar structural features, release intracellular Ca2+ from the IP3-sensitive pool, use different components in their signal transduction pathways in Xenopus oocytes.
Noh
S J
SJ
Department of Life Science, Pohang University of Science and Technology, South Korea.
Kim
M J
MJ
Shim
S
S
Han
J K
JK
D000818:Animals; D016375:Antisense Elements (Genetics); D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D002712:Chlorides / Q000378:metabolism; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D005260:Female; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000235:genetics / Q000378:metabolism; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000235:genetics / Q000378:metabolism; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism*; D008246:Lysophospholipids / Q000494:pharmacology*; D009544:Niflumic Acid / Q000494:pharmacology; D015345:Oligonucleotide Probes; D009865:Oocytes / Q000737:chemistry / Q000201:enzymology; D018408:Patch-Clamp Techniques; D010507:Periodicity; D054799:Phospholipase C beta; D011760:Pyrrolidinones / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D011956:Receptors, Cell Surface / Q000502:physiology*; D043562:Receptors, G-Protein-Coupled; D049312:Receptors, Lysophospholipid; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism*; D014982:Xenopus laevis
D016375:Antisense Elements (Genetics); D002614:Chelating Agents; D002712:Chlorides; D016861:Cyclooxygenase Inhibitors; D004791:Enzyme Inhibitors; D004963:Estrenes; D007527:Isoenzymes; D008246:Lysophospholipids; D015345:Oligonucleotide Probes; D011760:Pyrrolidinones; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D049312:Receptors, Lysophospholipid; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; C060506:sphingosine 1-phosphate; D009544:Niflumic Acid; D004533:Egtazic Acid; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D010738:Type C Phospholipases; D054799:Phospholipase C beta; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D019205:GTP-Binding Protein alpha Subunits, Gs; D013110:Sphingosine; D002118:Calcium