Diacylglycerol kinase epsilon, but not zeta selectively removes polyunsaturated diacylglycerol, inducing altered protein kinase C distribution in vivo

Porcine aortic endothelial cells have previously been shown to contain part icularly high basal levels of polyunsaturated diacylglycerol (DAG) together with a very high degree of membrane associated protein kinase C (PBC), whi ch is largely insensitive to further activation (Pettitt, T. R., Martin, A. Horton, T., Liossis, C., Lord, J. M., and Wakelam, M. J. O. (1997) J. Biol . Chem. 272, 17354-17359), To investigate the possibility that the high pol yunsaturated DAG levels were constitutively activating PKC, we transfected porcine aortic endothelial cells with two different forms of human diacylgl ycerol kinase, epsilon and zeta, In vitro, the former is specific for polyu nsaturated structures, whereas the latter shows no apparent selectivity, Ov erexpression of DAGK epsilon specifically reduced the level of polyunsatura ted DAG in the transfected cells while having little effect on the more sat urated structures. It also caused the redistribution of PKC alpha and epsil on from the membrane to the cytosol, Overexpression of DAGK zeta caused a g eneral reduction in DAG levels but had little effect on PKC distribution. T hese results for the first time show that DAGK epsilon specifically phospho rylates polyunsaturated DAG in vivo and that in so doing it regulates PKC l ocalization and activity, This provides support for the proposal that it is the polyunsaturated DAGs that function as messengers and convincing eviden ce for DAGK epsilon being a physiological terminator of DAG second messenge r signaling.