Ceramide regulates the transcription of cyclooxygenase-2. Evidence for involvement of extracellular signal-regulated kinase/c-Jun N-terminal kinase and p38 mitogen-activated protein kinase pathways.
The ceramide signaling pathway is activated by the sphingomyelinase (SMase)-mediated hydrolysis of cell membrane sphingomyelin to ceramide. We determined whether ceramide, a lipid second messenger, induced cyclooxygenase-2 (COX-2) in human mammary epithelial cells. Treatment of cells with neutral SMase or C2- or C6-ceramide enhanced prostaglandin E2 synthesis and increased levels of COX-2 protein and mRNA. Nuclear runoff assays revealed increased rates of COX-2 transcription after treatment with SMase and C2- and C6-ceramide. Transient transfections utilizing COX-2 promoter deletion constructs and COX-2 promoter constructs in which specific enhancer elements were mutagenized indicated that the effects of ceramide were mediated via a cAMP response element. The induction of COX-2 by ceramide was inhibited by calphostin C, an inhibitor of protein kinase C. Induction of COX-2 promoter activity by SMase was blocked by overexpressing kinase-deficient Raf-1. Triggering of the ceramide pathway also led to increases in extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (MAPK) activities; pharmacological inhibitors of MAPK kinase (MEK) and p38 MAPK blocked the induction of COX-2 by SMase. Overexpressing ERK1, JNK, or p38 led to severalfold increases in COX-2 promoter activity. By comparison, overexpression of dominant negatives for ERK1/2, JNK, or p38 blocked the activation of COX-2 promoter activity by SMase. A dominant negative for c-Jun inhibited the activation of COX-2 promoter activity by ceramide. Thus, in response to ceramide, increased MAPK signaling activates c-Jun, which, in turn, induces COX-2 gene expression via the cAMP response element in the COX-2 promoter.
Subbaramaiah
K
K
Department of Medicine, New York Hospital-Cornell Medical Center and Strang Cancer Prevention Center, New York, New York 10021, USA.
Chung
W J
WJ
Dannenberg
A J
AJ
D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics*; D002460:Cell Line; D002518:Ceramides / Q000502:physiology*; D051546:Cyclooxygenase 2; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D006801:Humans; D007527:Isoenzymes / Q000235:genetics*; D048031:JNK Mitogen-Activated Protein Kinases; D008565:Membrane Proteins; D020928:Mitogen-Activated Protein Kinases; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics*; D011453:Prostaglandins / Q000096:biosynthesis; D014158:Transcription, Genetic / Q000502:physiology*; D048051:p38 Mitogen-Activated Protein Kinases
D002518:Ceramides; D007527:Isoenzymes; D008565:Membrane Proteins; D011453:Prostaglandins; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases