Phospholipase C-gamma mediates the hydrolysis of phosphatidylinositol, but not of phosphatidylinositol 4,5-bisphoshate, in carbamylcholine-stimulated islets of langerhans.
In pancreatic islets the activation of phospholipase C (PLC) by the muscarinic receptor agonist carbamyolcholine (carbachol) results in the hydrolysis of both phosphatidylinositol 4,5-bisphosphate (PtdInsP(2)) and phosphatidylinositol (PtdIns). Here we tested the hypothesis that PtdIns hydrolysis is mediated by PLCgamma1, which is known to be regulated by activation of tyrosine kinases and PtdIns 3-kinase. PtdIns breakdown was more sensitive than that of PtdInsP(2) to the tyrosine kinase inhibitor, genistein. Conversely, the tyrosine phosphatase inhibitor, vanadate, alone promoted PtdIns hydrolysis and acted non-additively with carbachol. Vanadate did not stimulate PtdInsP(2) breakdown. Carbachol also stimulated a rapid (maximal at 1-2 min) tyrosine phosphorylation of several islet proteins, although not of PLCgamma1 itself. Two structurally unrelated inhibitors of PtdIns 3-kinase, wortmannin and LY294002, more effectively attenuated the hyrolysis of PtdIns compared with PtdInsP(2). Adenovirally mediated overexpression of PLCgamma1 significantly increased carbachol-stimulated PtdIns hydrolysis without affecting that of PtdInsP(2). Conversely overexpression of PLCbeta1 up-regulated the PtdInsP(2), but not PtdIns, response. These results indicate that the hydrolysis of PtdIns and PtdInsP(2) are independently regulated in pancreatic islets and that PLCgamma1 selectively mediates the breakdown of PtdIns. The activation mechanism of PLCgamma involves tyrosine phosphorylation (but not of PLCgamma directly) and PtdIns 3-kinase. Our findings point to a novel bifurcation of signaling pathways downstream of muscarinic receptors and suggest that hydrolysis of PtdIns and PtdInsP(2) might serve different physiological ends.
Mitchell
C J
CJ
Garvan Institute of Medical Research, St. Vincent's Hospital, Sydney 2010, Australia.
Kelly
M M
MM
Blewitt
M
M
Wilson
J R
JR
Biden
T J
TJ
D000256:Adenoviridae / Q000235:genetics; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002217:Carbachol / Q000494:pharmacology*; D018679:Cholinergic Agonists / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D002867:Chromones / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019833:Genistein / Q000494:pharmacology; D006868:Hydrolysis; D007295:Inositol Phosphates / Q000378:metabolism; D007515:Islets of Langerhans / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism* / Q000502:physiology*; D008564:Membrane Potentials; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism*; D010716:Phosphatidylinositols / Q000378:metabolism*; D051966:Phospholipase C gamma; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction; D013997:Time Factors; D010738:Type C Phospholipases / Q000378:metabolism* / Q000502:physiology*; D014443:Tyrosine / Q000378:metabolism; D014638:Vanadates / Q000494:pharmacology; D000077191:Wortmannin
D000730:Androstadienes; D018679:Cholinergic Agonists; D002867:Chromones; D004791:Enzyme Inhibitors; D007295:Inositol Phosphates; D007527:Isoenzymes; D009025:Morpholines; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D014638:Vanadates; D014443:Tyrosine; D002217:Carbachol; D019833:Genistein; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D000077191:Wortmannin