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Activation of protein kinase C family members by the novel polyphosphoinositides PtdIns-3,4-P2 and PtdIns-3,4,5-P3.

1994; Elsevier BV; Volume: 269; Issue: 51 Linguagem: Inglês

10.1016/s0021-9258(18)31643-0

1083-351X

Alex Toker, Maria Meyer, K. Kishta Reddy, John R. Falck, R. Aneja, Sarla G. Aneja, Alejandro Parra, David J. Burns, Lawrence M. Ballas, Lewis C. Cantley,

Melanoma and MAPK Pathways

The effect of phosphoinositides on the activity of protein kinase C (PKC) isotypes was investigated, PKC a, PI, PII, y, 6, E, 4, and 5 were expressed in baculovirusinfected insect cells and purified by column chromatography.The calcium-activated PKC isotypes a, PI, PII, and y were not significantly activated by any of the phosphoinositides investigated (phosphatidylinosito1-4phosphate (PtdTns-4-P), PtdIns-3-P, PtdIns-4,5-P,, Pt-dIns-3,4-P2, and PtdIns-3,4,5-P3) when added in the presence of concentrations of phosphatidylserine that give maximal stimulation.The calcium-insensitive PKC isotypes 6, E, and 9 also showed little response to PtdIns-3-P, PtdIns4-P, or PtdIns-4,5-P2 when these lipids were added in the presence of phosphatidylserine.In contrast, PtdIns-3,4-PZ and PtdIns-3,4,5-P3 caused a 5-15fold stimulation of these enzymes compared with phosphatidylserine alone.50 % maximal stimulation of PKC E by PtdIns-3,4,5-P3 occurred when this lipid was present at about 1% of the carrier PtdIns-4,5-P2 (about 100 m).These lipids had little effect on baculovirus-expressed PKC f which was constitutively active.A short chain version of PtdIns-3,4,5-P3, dioctanoyl-PtdIns-3,4,5-P3, activated PKC 6, E , and 4 in the absence of other lipids, whereas a short chain version of PtdIns-4,5-P,, dihexanoyl-PtdIns-4,5-P2, did not.Since PtdIns-3,4-P2 and Pt-dIns-3,4,5-P, are nominally absent in unstimulated cells and appear within seconds to minutes of stimulation by various cell activators, these lipids could act as second messengers to activate PKC 6, E, or q in vivo.Phosphoinositide 3-kinase is stimulated by a host of growth factors and other cellular activators (1).In vitro, this enzyme can phosphorylate phosphatidylinositol (PtdIns),' PtdIns-4-P, and A. T.) and GM37361 and GM49594 (to R. A,).The costs of publica-