Cloning and Characterization of a Glucocorticoid-induced Diacylglycerol Kinase

Abstract

Diacylglycerol kinase (DGK) plays a key role in cellular processes by regulating the intracellular concentration of the second messenger diacylglycerol. We screened a hamster DDT1 smooth muscle cell library and isolated a unique, glucocorticoid-inducible cDNA with substantial homology to known DGKs. DGK activity was increased in lysates of insect cells infected with recombinant baculovirus containing this cDNA. Antibodies raised against expressed sequences recognized a glucocorticoid-inducible 130-140-kDa protein on immunoblots of DDT1 cell lysates. Thus, this sequence appears to be a new member of the DGK family that we refer to as DGKeta. Homology to other DGKs was apparent in domains that are thought to be important for DGK function including the cysteine-rich motifs and potential catalytic domains. DGKeta shares substantial homology with DGKdelta including the N-terminal pleckstrin homology domain. The tissue distribution of DGKeta message (determined by ribonuclease protection assays) and protein (determined by immunoblots) was broader than reported for other DGKs, indicating that DGKeta may play a more general role in regulating cellular DG levels than other DGKs. Heterogeneity among DGK family members indicates that individual DGKs may have unique functions.