Ca2+ regulates the inositol tris/tetrakisphosphate pathway in intact and broken preparations of insulin-secreting RINm5F cells.
The two-step isomerization of inositol 1,4,5-trisphosphate (Ins-1,4,5-P3) to Ins-1,3,4-P3 via the intermediate inositol 1,3,4,5-tetrakisphosphate (Ins-P4) was studied in intact RINm5F cells and in subcellular fractions. Muscarinic stimulation with carbamylcholine leads to a rapid (2 s) rise in both Ins-1,4,5-P3 and Ins-P4, whereas Ins-1,3,4-P3 was produced only after a lag of at least 5 s. In cells with depleted Ca2+ stores, the rise in Ins-1,4,5-P3 was nearly tripled, and that of Ins-1,3,4-P3 markedly diminished as compared to control cells. Raising the free Ca2+ concentration from 10(-7) to 10(-5) M increased inositol 1,4,5-triphosphate-3-kinase activity in cytosolic fractions by 2 1/2-fold (EC50 for Ca2+ approximately 0.8 microM) but had no effect on the activity of inositol 1,4,5-triphosphate-5-phosphomonoesterase. At 10(-7) M Ca2+ these two enzymes displayed comparable activity when assayed at concentrations of Ins-1,4,5-P3 occurring in stimulated cells; however, at 10(-5) M Ca2+, kinase activity predominates. These results suggest that Ins-1,4,5-P3 counter-regulates its own levels through the activity of inositol 1,4,5-trisphosphate 3-kinase and that the increase in [Ca2+]i may account for the transience of the rise in Ins-1,4,5-P3 seen during muscarinic stimulation of RINm5F cells.
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D015544:Inositol 1,4,5-Trisphosphate; D007295:Inositol Phosphates / Q000378:metabolism*; D000072181:Inositol Polyphosphate 5-Phosphatases; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010770:Phosphotransferases / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor); D011494:Protein Kinases / Q000378:metabolism; D013403:Sugar Phosphates / Q000378:metabolism*
D007295:Inositol Phosphates; D007328:Insulin; D013403:Sugar Phosphates; C081655:inositol-1,3,4,5-tetrakisphosphate; D015544:Inositol 1,4,5-Trisphosphate; D002217:Carbachol; D010770:Phosphotransferases; D011494:Protein Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); C051038:Inositol 1,4,5-trisphosphate 3-kinase; D010744:Phosphoric Monoester Hydrolases; D000072181:Inositol Polyphosphate 5-Phosphatases; D002118:Calcium