Transcriptional regulation of the human neutral ceramidase gene

Sean M. O'Neill, Jong Yun, Todd E. Fox, Mark Kester

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Ceramidases play a critical role in generating sphingosine-1-phosphate by hydrolyzing ceramide into sphingosine, a substrate for sphingosine kinase. In order to elucidate its transcriptional regulation, we identify here a putative promoter region in the 5′-UTR of the human neutral CDase (nCDase) gene. Using human genomic DNA, we cloned a 3000 bp region upstream of the translational start site of the nCDase gene. Luciferase reporter analyses demonstrated that this 3000 bp region had promoter activity, with the strongest induction occurring within the first 200 bp. Computational analysis revealed the 200 bp essential promoter region contained several well-characterized promoter elements, lacked a conical TATA box, but did contain a reverse oriented CCAAT box, a feature common to housekeeping genes. Electrophoretic mobility shift assays demonstrated that the identified candidate transcriptional response elements (TRE) bind their respective transcription factors, including NF-Y, AP-2, Oct-1, and GATA. Mutagenic analyses of the TRE revealed that these sites regulated promoter activity and mutating an individual site decreased promoter reporter activity by up to 50%. Together, our findings suggest that regulation of nCDase expression involves coordinated TATA-less transcriptional activity.

Original languageEnglish (US)
Pages (from-to)21-30
Number of pages10
JournalArchives of Biochemistry and Biophysics
Volume511
Issue number1-2
DOIs
StatePublished - Jul 1 2011

Fingerprint

Genetic Promoter Regions
Genes
Response Elements
Ceramidases
Electrophoretic mobility
TATA Box
Sphingosine
Ceramides
5' Untranslated Regions
Essential Genes
Electrophoretic Mobility Shift Assay
Luciferases
Assays
Transcription Factors
DNA
Substrates
human ASAH2 protein

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology

Cite this

O'Neill, Sean M. ; Yun, Jong ; Fox, Todd E. ; Kester, Mark. / Transcriptional regulation of the human neutral ceramidase gene. In: Archives of Biochemistry and Biophysics. 2011 ; Vol. 511, No. 1-2. pp. 21-30.
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Transcriptional regulation of the human neutral ceramidase gene. / O'Neill, Sean M.; Yun, Jong; Fox, Todd E.; Kester, Mark.

In: Archives of Biochemistry and Biophysics, Vol. 511, No. 1-2, 01.07.2011, p. 21-30.

Research output: Contribution to journalArticle

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