title: AP-1 binding transcriptionally regulates human neutral ceramidase.
abstract: Many forms of cellular stress cause an elevation of endogenous ceramide levels leading to growth arrest or apoptosis. Ceramidases (CDase) play a critical role in regulating apoptosis by hydrolyzing ceramide into sphingosine, a precursor for promitogenic sphingosine-1-phosphate. Growth factor induction of neutral CDase (nCDase) has been shown to have a cytoprotective effect against cytokine-induced increases in ceramide levels. To further define the physiological regulation of nCDase, we identified a 200 bp promoter region and demonstrated that serum activated this proximal promoter, which correlated with a serum-induced increase in human nCDase mRNA expression. Computational analysis revealed a putative cis-element for AP-1, a transcription factor activated by serum. Electrophoretic mobility shift assays demonstrated that the identified transcriptional response element binds to AP-1 transcription factors. RNA interference-mediated knockdown of the AP-1 subunit, c-Jun, inhibited the activity of the human nCDase proximal promoter, whereas, c-Jun overexpression increased promoter activity, which directly correlated with human nCDase mRNA transcription, decreased ceramide mass, and protection against caspase 3/7-dependent apoptosis. Taken together, our findings suggest that c-Jun/AP-1 signaling may, in part, regulate serum-induced human nCDase gene transcription.
authors.affiliation: Department of Pharmacology, Penn State College of Medicine, Hershey, PA 17033, USA.
authors.forename: Sean M
authors.initials: SM
authors.lastname: O'Neill
authors.forename: Kristy L
authors.initials: KL
authors.lastname: Houck
authors.forename: Jong K
authors.initials: JK
authors.lastname: Yun
authors.forename: Todd E
authors.initials: TE
authors.lastname: Fox
authors.forename: Mark
authors.initials: M
authors.lastname: Kester
mesh_terms: D017209:Apoptosis / Q000235:genetics / Q000502:physiology; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002518:Ceramides / Q000378:metabolism; D003470:Culture Media; D017931:DNA Primers / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D055785:Gene Knockdown Techniques; D016762:Genes, fos; D057809:HEK293 Cells; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics; D055576:Neutral Ceramidase / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D050436:Regulatory Elements, Transcriptional; D015398:Signal Transduction; D018808:Transcription Factor AP-1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D002518:Ceramides; D003470:Culture Media; D017931:DNA Primers; D012333:RNA, Messenger; D018808:Transcription Factor AP-1; D048031:JNK Mitogen-Activated Protein Kinases; C526744:ASAH2 protein, human; D055576:Neutral Ceramidase