Regulation of rat ornithine decarboxylase promoter activity by binding of transcription factor Sp1

A. P. Kumar, P. K. Mar, B. Zhao, R. L. Montgomery, D. C. Kang, A. P. Butler

Research output: Contribution to journalArticle

44 Scopus citations

Abstract

Ornithine decarboxylase (ODC) is the rate-limiting enzyme of polyamine biosynthesis. We investigated the transcriptional regulation of the rat ODC gene using transient expression assays. The 5'-flanking region (-1156 to +13) of the ODC gene was sufficient to mediate strong basal expression of a luciferase reporter. Sequences between -345 and -93 contributed to basal promoter activity. This region, containing five potential Sp1 binding sites, was analyzed by electrophoretic mobility shift assays. Three specific DNA- protein complexes were identified using H35 nuclear extracts and the -345/- 93 ODC probe. Binding to all three was eliminated by competition with an oligonucleotide containing an Sp1 binding site, but not by a mutant Sp1 oligonucleotide. Preincubation with an antibody against Sp1 supershifted complexes associated with one or more of Sp1 binding sites 1-4 as well as with site 5. DNase I footprinting revealed two protected regions: PR-I (-92 to -130) and PR-II (-304 to -332). PR-I contains a putative binding site for Sp1 that was protected by recombinant Sp1 protein. Transfection studies in Schneider SL2 cells demonstrated that the ODC promoter is trans-activated up to 350-fold by Sp1 and that this trans-activation is dependent on the presence of Sp1 binding sites 1-4. Thus, although the ODC promoter binds multiple nuclear proteins, Sp1 or a related protein appears to be a critical determinant of ODC transcription, possibly through cooperative interactions between Sp1 and additional transcription factors.

Original languageEnglish (US)
Pages (from-to)4341-4348
Number of pages8
JournalJournal of Biological Chemistry
Volume270
Issue number9
DOIs
StatePublished - Jan 1 1995
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Fingerprint Dive into the research topics of 'Regulation of rat ornithine decarboxylase promoter activity by binding of transcription factor Sp1'. Together they form a unique fingerprint.

  • Cite this