Expression of functional receptor-coupled TRPC3 channels in DT40 triple receptor InsP3 knockout cells.
The TRPC3 channel, an intensively studied member of the widely expressed transient receptor potential (TRP) family, is a Ca(2+)-conducting channel activated in response to phospholipase C-coupled receptors. Despite scrutiny, the receptor-induced mechanism to activate TRPC3 channels remains unclear. Evidence indicates TRPC3 channels interact directly with intracellular inositol 1,4,5-trisphosphate receptors (InsP(3)Rs) and that channel activation is mediated through coupling to InsP(3)Rs. TRPC3 channels were expressed in DT40 chicken B lymphocytes in which all three InsP(3)R genes were deleted (DT40InsP(3)R-k/o). Endogenous B-cell receptors (BCR) coupled through Syk kinase to phospholipase C-gamma (PLC-gamma) activated the expressed TRPC3 channels in both DT40w/t and DT40InsP(3)R-k/o cells. The diacylglycerol (DAG) analogue 1-oleoyl-2-acetyl-sn-glycerol (OAG) also activated TRPC3 channels independently of InsP(3)Rs. BCR-induced TRPC3 activation was blocked by the PLC enzymic inhibitor, U-73122, and also blocked by wortmannin-induced PLC substrate depletion. Neither U-73122 nor wortmannin modified either OAG-induced TRPC3 activation or store-operated channel activation in DT40 cells. Cotransfection of cells with both G protein-coupled M5 muscarinic receptors and TRPC3 channels resulted in successful M5 coupling to open TRPC3 channels mediated by PLC-beta. We conclude that TRPC3 channels are activated independently of InsP(3)Rs through DAG production resulting from receptor-mediated activation of either PLC-gamma or PLC-beta.
Venkatachalam
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Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
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Ford
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Gill
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D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D030801:Animals, Genetically Modified; D001402:B-Lymphocytes / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000235:genetics* / Q000502:physiology; D002460:Cell Line; D002478:Cells, Cultured; D002645:Chickens; D018076:DNA, Complementary / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004792:Enzyme Precursors / Q000378:metabolism; D004963:Estrenes / Q000494:pharmacology; D053496:Inositol 1,4,5-Trisphosphate Receptors; D047908:Intracellular Signaling Peptides and Proteins; D007473:Ion Channels / Q000096:biosynthesis*; D007527:Isoenzymes / Q000378:metabolism; D051379:Mice; D009154:Mutation; D051966:Phospholipase C gamma; D010957:Plasmids / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011760:Pyrrolidinones / Q000494:pharmacology; D011947:Receptors, Antigen, B-Cell / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics* / Q000502:physiology; D000072377:Syk Kinase; D050052:TRPC Cation Channels; D013997:Time Factors; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism; D000077191:Wortmannin
D000730:Androstadienes; D015220:Calcium Channels; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D004792:Enzyme Precursors; D004963:Estrenes; D053496:Inositol 1,4,5-Trisphosphate Receptors; D047908:Intracellular Signaling Peptides and Proteins; D007473:Ion Channels; D007527:Isoenzymes; D011760:Pyrrolidinones; D011947:Receptors, Antigen, B-Cell; D018160:Receptors, Cytoplasmic and Nuclear; D050052:TRPC Cation Channels; C104603:TRPC3 cation channel; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D011505:Protein-Tyrosine Kinases; D000072377:Syk Kinase; C000606897:Syk protein, mouse; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D002118:Calcium; D000077191:Wortmannin