Inositol 1,4,5 trisphosphate is inactivated by a 5-phosphatase in stamen hair cells of Tradescantia

Inositol 1,4,5 trisphosphate [Ins(1,4,5)P-3] is produced from the hydrolysi s of phosphatidylinositol 4,5 bisphosphate, and as part of a second-messeng er signal transduction mechanism, induces release of Ca2+ from internal sto res in both plant and animal systems. It is less well established how the a ctive Ins(1,4,5)P-3 is inactivated. Studies in animal cells have demonstrat ed two separate metabolic pathways. Ins(1,4,5)P-3 can be hydrolyzed by a 5- phosphatase or phosphorylated by a 3-kinase, resulting in the formation of Ins(1,4)P-2 and Ins(1,3,4,5)P-4, respectively, neither of which is able to mobilize intracellular Ca2+. Plant cell extracts have been reported to have hydrolytic and kinase activities that produce Ins(1,4)P-2, and Ins(4,5)P-2 and Ins(1,4,5,6)P-4 from Ins(1,4,5)P-3. These results offer little insight into the enzyme activities in the intact plant cell since the observed act ivities might be confined to intracellular compartments that have little if any impact on the signaling events within the cytosol that require Ins(1,4 ,5)P-3. To resolve the mechanism of Ins(1,4,5)P-3 inactivation, we microinj ected stamen hair cells of Tradescantia virginiana L. with nonhydrolysable analogs of Ins(1,4,5)P-3 that have been previously shown to cause Ca2+ rele ase from intracellular stores. Our results indicate a sustained cytosolic [ Ca2+] increase when cells were injected with the 5-phosphatase-insensitive 5-monophosphorothioate derivative of Ins(1,4,5)P-3, in contrast to a brief transient when injected with the 3-kinase-insensitive 3-fluoro-3-deoxy Ins( 1,4,5)P-3 analog. We conclude that the 5-phosphatase pathway is the preferr ed pathway for Ins(1,4,5)P-3 inactivation in the stamen hair cells of Trade scantia.