The synthesis of inositol polyphosphates in rat liver by 5- and 6-kinase activities.

Amongst the array of known inositol phosphates, Ins(1,3,4)P3 is a pivotal intermediate. It can be attacked by two distinct phosphatases, and can also be phosphorylated at the 5- and 6-positions. The kinases that perform the latter reactions co-purified during anion-exchange chromatography; the 5-kinase accounted for 16% of the total kinase activity. The two resultant InsP4 isomers were identified by periodate oxidation, alkaline hydrolysis and specific enzyme assays. The phosphorylation of Ins(1,3,4)P3 to Ins(1,3,4,5)P4 may be important in coordinating calcium signalling. The production of Ins(1,3,4,6)P4 may provide a link between receptor-activated inositol phosphate production and the synthesis of InsP5 and InsP6, because Ins(1,3,4,6)P4 can itself be further phosphorylated. Two peaks of Ins(1,3,4,6)P4 kinase activity were separated by anion-exchange chromatography. One of these was a 5-kinase, and the other appeared to be a mixture of 5-kinase and 2-kinase activities. We investigated how the metabolism of Ins(1,3,4)P3 might be controlled in rat liver supernatant, which contained most of the hepatic Ins(1,3,4)P3 phosphatase and kinase activities. Ins(1,3,4)P3 phosphorylation was subject to kinetic restraints, such that the dephosphorylation pathway was greatly favoured. Moreover Ins(3,4,5,6)P4, which is itself an alternative precursor of InsP5, was a potent inhibitor of Ins(1,3,4)P3 kinase. We do not yet know the concentration of Ins(3,4,5,6)P4 in liver, although indirect evidence supports the idea that it is an endogenous inhibitor of Ins(1,3,4)P3 kinase. Our results illustrate the relationship between receptor-mediated inositol phosphate production and the synthesis of higher polyphosphates in rat liver, whilst also providing some insight into how this metabolic association is controlled.