CerS6 Is a Novel Transcriptional Target of p53 Protein Activated by Non-genotoxic Stress.
Our previous study suggested that ceramide synthase 6 (CerS6), an enzyme in sphingolipid biosynthesis, is regulated by p53: CerS6 was elevated in several cell lines in response to transient expression of p53 or in response to folate stress, which is known to activate p53. It was not clear, however, whether CerS6 gene is a direct transcriptional target of p53 or whether this was an indirect effect through additional regulatory factors. In the present study, we have shown that the CerS6 promoter is activated by p53 in luciferase assays, whereas transcriptionally inactive R175H p53 mutant failed to induce the luciferase expression from this promoter. In vitro immunoprecipitation assays and gel shift analyses have further demonstrated that purified p53 binds within the CerS6 promoter sequence spanning 91 bp upstream and 60 bp downstream of the transcription start site. The Promo 3.0.2 online tool for the prediction of transcription factor binding sites indicated the presence of numerous putative non-canonical p53 binding motifs in the CerS6 promoter. Luciferase assays and gel shift analysis have identified a single motif upstream of the transcription start as a key p53 response element. Treatment of cells with Nutlin-3 or low concentrations of actinomycin D resulted in a strong elevation of CerS6 mRNA and protein, thus demonstrating that CerS6 is a component of the non-genotoxic p53-dependent cellular stress response. This study has shown that by direct transcriptional activation of CerS6, p53 can regulate specific ceramide biosynthesis, which contributes to the pro-apoptotic cellular response.
From the Nutrition Research Institute, University of North Carolina at Chapel Hill, Kannapolis, North Carolina 28081.
Baharan
B
Fekry
From the Nutrition Research Institute, University of North Carolina at Chapel Hill, Kannapolis, North Carolina 28081.
Kristen A
KA
Jeffries
From the Nutrition Research Institute, University of North Carolina at Chapel Hill, Kannapolis, North Carolina 28081.
Amin
A
Esmaeilniakooshkghazi
the Department of Biochemistry and Hollings Cancer Center, Medical University of South Carolina, Charleston, South Carolina 29425, and.
Besim
B
Ogretmen
From the Nutrition Research Institute, University of North Carolina at Chapel Hill, Kannapolis, North Carolina 28081, the Department of Nutrition, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599.
Sergey A
SA
Krupenko
From the Nutrition Research Institute, University of North Carolina at Chapel Hill, Kannapolis, North Carolina 28081, the Department of Nutrition, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599 Natalia_krupenko@unc.edu.
Natalia I
NI
Krupenko
D000072283:A549 Cells; D019943:Amino Acid Substitution; D002518:Ceramides / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D020125:Mutation, Missense; D059372:Nucleotide Motifs; D010879:Piperazines / Q000494:pharmacology; D020218:Response Elements; D051049:Sphingosine N-Acyltransferase / Q000235:genetics / Q000378:metabolism*; D013312:Stress, Physiological; D014158:Transcription, Genetic; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*
D002518:Ceramides; D007093:Imidazoles; D008565:Membrane Proteins; D010879:Piperazines; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; C482205:nutlin 3; C506123:CERS6 protein, human; D051049:Sphingosine N-Acyltransferase
ceramide synthase; lipid signaling; p53; transcription factor; transcription regulation