Interaction of the Type I alpha PIPkinase with phospholipase D: a role forthe local generation of phosphatidylinositol 4,5-bisphosphate in the regulation of PLD2 activity

Phosphoinositides are localized in various intracellular compartments and c an regulate a number of intracellular functions, such as cytoskeletal dynam ics and membrane trafficking. Phospholipase Ds (PLDs) are regulated enzymes that hydrolyse phosphatidylcholine (PtdCho) to generate the putative secon d messenger phosphatidic acid (PtdOH). In vitro, PLDs have an absolute requ irement for higher phosphorylated inositides, such as phosphatidylinositol 4,5-bis-phosphate [PtdIns(4,5)P-2]. Whether this lipid is able to regulate the activity of PLD in vivo is contentious. To examine this hypothesis we s tudied the relationship between PLD and an enzyme critical for the intracel lular synthesis of PtdIns(4,5)P-2: phosphatidylinositol 4-phosphate 5-kinas e alpha (Type I alpha PIPkinase). We find that both PLD1 and PLD2 interact with the Type I alpha PIPkinase and that PLD2 activity in vivo can be regul ated solely by the expression of this lipid kinase. Moreover, PLD2 is able to recruit the Type I alpha PIPkinase to its intracellular location. We sho w that the physiological requirement of PLD enzymes for PtdIns(4,5)P-2 is c ritical and that PLD2 activity can be regulated solely by the levels of thi s key intracellular lipid.