Alternative splicing of the TRPC3 ion channel calmodulin/IP3 receptor-binding domain in the hindbrain enhances cation flux.
Canonical transient receptor potential (TRPC3) nonselective cation channels are effectors of G-protein-coupled receptors (GPCRs), activated via phospholipase C-diacylglycerol signaling. In cerebellar Purkinje cells, TRPC3 channels cause the metabotropic glutamate receptor (mGluR)-mediated slow EPSC (sEPSC). TRPC3 channels also provide negative feedback regulation of cytosolic Ca(2+), mediated by a C terminus "calmodulin and inositol trisphosphate receptor binding" (CIRB) domain. Here we report the alternative splicing of the TRPC3 mRNA transcript (designated TRPC3c), resulting in omission of exon 9 (approximately half of the CIRB domain) in mice, rats, and guinea pigs. TRPC3c expression is brain region specific, with prevalence in the cerebellum and brainstem. The TRPC3c channels expressed in HEK293 cells exhibit increased basal and GPCR-activated channel currents, and increased Ca(2+) fluorescence responses, compared with the previously characterized (TRPC3b) isoform when activated via either the endogenous M3 muscarinic acetylcholine receptor, or via coexpressed mGluR1. GPCR-induced TRPC3c channel opening rate (cell-attached patch) matched the maximum activation achieved with inside-out patches with zero cytosolic Ca(2+), whereas the GPCR-induced TRPC3b activation frequency was significantly less. Both TRPC3 channel isoforms were blocked with 2 mm Ca(2+), attributable to CIRB domain regulation. In addition, genistein blocked Purkinje cell (S)-2-amino-2-(3,5-dihydroxyphenyl) acetic acid (mGluR1)-activated TPRC3 current as for recombinant TRPC3c current. This novel TRPC3c ion channel therefore has enhanced efficacy as a neuronal GPCR-Ca(2+) signaling effector, and is associated with sensorimotor coordination, neuronal development, and brain injury.
Translational Neuroscience Facility and Department of Physiology, School of Medical Sciences, The University of New South Wales, Sydney, New South Wales 2052, Australia.
Youngsoo
Y
Kim
Ann Chi Yan
AC
Wong
John M
JM
Power
Sherif F
SF
Tadros
Matthias
M
Klugmann
Andrew J
AJ
Moorhouse
Paul P
PP
Bertrand
Gary D
GD
Housley
D017398:Alternative Splicing / Q000235:genetics*; D000595:Amino Acid Sequence; D000704:Analysis of Variance; D000818:Animals; D055592:Biophysical Phenomena / Q000187:drug effects / Q000235:genetics; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000235:genetics / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology; D002461:Cell Line, Transformed; D002531:Cerebellum / Q000166:cytology*; D018679:Cholinergic Agonists / Q000494:pharmacology; D018683:Excitatory Amino Acid Agents / Q000494:pharmacology; D005260:Female; D019833:Genistein / Q000494:pharmacology; D006168:Guinea Pigs; D006801:Humans; D053496:Inositol 1,4,5-Trisphosphate Receptors / Q000235:genetics / Q000378:metabolism*; D008297:Male; D008734:Methoxyhydroxyphenylglycol / Q000031:analogs & derivatives / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051381:Rats; D050052:TRPC Cation Channels / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D002147:Calmodulin; D018679:Cholinergic Agonists; D018683:Excitatory Amino Acid Agents; D053496:Inositol 1,4,5-Trisphosphate Receptors; D020033:Protein Isoforms; D047428:Protein Kinase Inhibitors; D050052:TRPC Cation Channels; C104603:TRPC3 cation channel; D008734:Methoxyhydroxyphenylglycol; D002217:Carbachol; D019833:Genistein; D002118:Calcium; C010117:3,4-dihydroxyphenylglycol