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 particularly high basal levels of polyunsaturated diacylglycerol (DAG) together with a very high degree of membrane-associated protein kinase C (PKC), which 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 polyunsaturated DAG levels were constitutively activating PKC, we transfected porcine aortic endothelial cells with two different forms of human diacylglycerol kinase, epsilon and zeta. In vitro, the former is specific for polyunsaturated structures, whereas the latter shows no apparent selectivity. Overexpression of DAGKepsilon specifically reduced the level of polyunsaturated DAG in the transfected cells while having little effect on the more saturated structures. It also caused the redistribution of PKCalpha and epsilon from the membrane to the cytosol. Overexpression of DAGKzeta caused a general reduction in DAG levels but had little effect on PKC distribution. These results for the first time show that DAGKepsilon specifically phosphorylates polyunsaturated DAG in vivo and that in so doing it regulates PKC localization and activity. This provides support for the proposal that it is the polyunsaturated DAGs that function as messengers and convincing evidence for DAGKepsilon being a physiological terminator of DAG second messenger signaling.
Pettitt
T R
TR
Institute for Cancer Studies, The University of Birmingham, Birmingham B15 2TA, United Kingdom.
Wakelam
M J
MJ
D019852:Diacylglycerol Kinase / Q000235:genetics / Q000378:metabolism*; D004075:Diglycerides / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D015398:Signal Transduction
D004075:Diglycerides; D007527:Isoenzymes; D019852:Diacylglycerol Kinase; D011493:Protein Kinase C