Characterization of the human UDP-galactose: Ceramide galactosyltransferase gene promoter

Tewin Tencomnao, Robert K Yu, Dmitri Kapitonov

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

UDP-galactose:ceramide galactosyltransferase (CGT, EC 2.4.1.45) is a key enzyme in the biosynthesis of galactocerebroside, the most abundant glycosphingolipid in the myelin sheath. An 8 kb fragment upstream from the transcription initiation site of CGT gene was isolated from a human genomic DNA library. Primer extension analysis revealed a single transcription initiation site 329 bp upstream from the ATG start codon. Neither a consensus TATA nor a CCAAT box was identified in the proximity to the transcription start site; however, this region contains a high GC content and multiple putative regulatory elements. To investigate the transcriptional regulation of CGT, a series of 5′ deletion constructs of the 5′-flanking region were generated and cloned upstream from the luciferase reporter gene. By comparing promoter activity in the human oligodendroglioma (HOG) and human neuroblastoma (LAN-5) cell lines, we found that the CGT promoter functions in a cell type-specific manner. Three positive cis-acting regulatory regions were identified, including a proximal region at -292/-256 which contains the potential binding sites for known transcription factors (TFs) such as Ets and SP1 (GC box), a distal region at -747/-688 comprising a number of binding sites such as the ERE half-site, NF1-like, TGGCA-BP, and CRE, and a third positive cis-acting region distally localized at -1325/-1083 consisting of binding sites for TFs such as nitrogen regulatory, TCF-1, TGGCA-BP, NF-IL6, CF1, bHLH, NF1-like, GATA, and γ-IRE. A negative cis-acting domain localized in a far distal region at -1594/-1326 was also identified. Our results suggest the presence of both positive and negative cis-regulatory regions essential for the cell-specific expression in the TATA-less promoter of the human CGT gene.

Original languageEnglish (US)
Pages (from-to)416-423
Number of pages8
JournalBiochimica et Biophysica Acta - Gene Structure and Expression
Volume1517
Issue number3
DOIs
StatePublished - Feb 16 2001

Fingerprint

Ganglioside Galactosyltransferase
Transcription Initiation Site
Genes
Nucleic Acid Regulatory Sequences
Binding Sites
Transcription Factors
Local Area Networks
CCAAT-Enhancer-Binding Protein-beta
Oligodendroglioma
Glycosphingolipids
Initiator Codon
Genomic Library
5' Flanking Region
Biosynthesis
Base Composition
Myelin Sheath
Luciferases
Neuroblastoma
Gene Library
Reporter Genes

Keywords

  • Gene expression
  • Myelination
  • Promoter
  • Transcription
  • Uridine diphosphate-galactose:ceramide galactosyltransferase

ASJC Scopus subject areas

  • Structural Biology
  • Biophysics
  • Biochemistry
  • Genetics

Cite this

Characterization of the human UDP-galactose : Ceramide galactosyltransferase gene promoter. / Tencomnao, Tewin; Yu, Robert K; Kapitonov, Dmitri.

In: Biochimica et Biophysica Acta - Gene Structure and Expression, Vol. 1517, No. 3, 16.02.2001, p. 416-423.

Research output: Contribution to journalArticle

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