Inositol 1:2(cyclic),4,5-trisphosphate is not a major product of inositol phospholipid metabolism in vasopressin-stimulated WRK1 cells.
1. A method has been devised for quenching cell incubations with an aqueous phenol/chloroform/EDTA mixture of neutral pH, to allow the analysis of acid-labile cell components. 2. Using this method, we have searched for the appearance of Ins(1:2cyclic,4,5)P3 [inositol 1:2(cyclic),4,5-trisphosphate] in WRK1 mammary tumour cells that were labelled to high specific radioactivity with [3H]inositol and then stimulated with 0.4 microM-vasopressin. 3. Vasopressin caused a very rapid accumulation of Ins(1,4,5)P3 (inositol 1,4,5-trisphosphate), followed by a slower decline towards the original concentration. An acid-labile and inositol-labelled compound with the chromatographic properties of Ins(1:2cyclic,4,5)P3 was present in unstimulated cells at less than 5% of the elevated concentration of Ins(1,4,5)P3. Its concentration rose 2-3-fold during stimulation for 3 min, at which time its concentration was about 5% of the elevated concentration of Ins(1,4,5)P3. 4. We conclude that Ins(1,4,5)P3 is the major product of phosphoinositidase C-catalysed phosphatidylinositol 4,5-bisphosphate hydrolysis in vasopressin-stimulated WRK1 cells. Ins(1:2cyclic,4,5)P3 is unlikely to be an important intracellular messenger in these cells, at least during the first few minutes of stimulation.
Department of Biochemistry, University of Birmingham, U.K.
N S
NS
Wong
C J
CJ
Barker
S B
SB
Shears
C J
CJ
Kirk
R H
RH
Michell
D001127:Arginine Vasopressin / Q000494:pharmacology*; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D007295:Inositol Phosphates / Q000378:metabolism*; D010716:Phosphatidylinositols / Q000378:metabolism*; D013403:Sugar Phosphates / Q000378:metabolism*
D007295:Inositol Phosphates; D010716:Phosphatidylinositols; D013403:Sugar Phosphates; D001127:Arginine Vasopressin; C049924:inositol-1,2-cyclic-4,5-triphosphate