Changes in phosphoinositide metabolism with days in culture affect signal transduction pathways in Galdieria sulphuraria

The metabolism of phosphatidylinositol-4,5-bisphosphate (PIP,) changed duri ng the culture period of the thermoacidophilic red alga Galdieria sulphurar ia. Seven days after inoculation, the amount of PIP2 in the cells was 910 /- 100 pmol g(-1) fresh weight; by 12 d, PIP2 levels increased to 1200 +/- 150 pmol g(-1) fresh weight. In vitro assays indicated that phosphatidylino sitol monophosphate (PIP) kinase specific activity increased from 75 to 230 pmol min(-1) mg(-1) protein between d 7 and 12. When G. sulphuraria cells were osmostimulated, transient increases of up to I-fold could be observed in inositol-1,4,5-trisphosphate (IP3) levels within 90 s, regardless of the age of the cells. In d-12 cells, the increase in IP3 was preceded by a tra nsient increase of up to 5-fold in specific PIP kinase activity, whereas no such increase was detected after osmo-stimulation of d-7 cells. The increa se in PIP kinase activity before IP3 signaling in d-12 cells indicates that there is an additional pathway for regulation of phosphoinositide metaboli sm after stimulation other than an initial activation of phospholipase C. A lso, the rapid activation of PIP2 biosynthesis in cells with already-high P IP, levels suggests that the PIP2 present was not available for signal tran sduction. By comparing the response of the cells at d 7 and 12, we have ide ntified two potentially distinct pools of PIP2.