The tert-butylhydroquinone-mediated activation of the human thioredoxin gene reveals a novel promoter structure

Simone A. Osborne, Hye-Jin Kim Hawkes, Ben L. Baldwin, Kylie A. Alexander, Terje Svingen, Frank M. Clarke, Kathryn F. Tonissen

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

Thioredoxin is a redox-active protein that plays multiple roles in regulating cell growth, cell signalling and apoptosis. Here, we have demonstrated that a complex mechanism involving multiple regulatory elements is involved in the tBHQ [tert-butylhydroquinone or 2,5-di-(t-butyl)-1,4-hydroquinonel-mediated activation of the thioredoxin gene. Luciferase assays, utilizing various wild-type and mutated thioredoxin promoter fragments, revealed roles for the ORE (oxidative stress responsive element), ARE (antioxidant responsive element), three Sp1 (specificity protein I)binding sites and the TATA box in the activation of the thioredoxin gene by tBHQ. The ORE required the presence of the ARE to elicit its response, whereas the independent removal of three Sp1-binding sites and the TATA box also decreased activation of the thioredoxin gene, with mutation of the TATA box having the greatest effect. Real-time RT (reverse transcriptase)-PCR analysis also revealed varying roles for two TSSs (transcription start sites) in the activation of the thioredoxin gene by tBHQ. Transcription was initiated from both TSSs; however, different response rates and fold inductions were observed. Together, these results suggest that the thioredoxin gene is controlled by a novel arrangement of two overlapping core promoter regions, one containing a TATA box and the other TATA-less. Altering the intracellular levels of thioredoxin in a breast cancer cell line also influenced the induction of thioredoxin transcription in response to tBHQ. Stable transfections with a redox-inactive thioredoxin mutant produced 3.6 times higher induction levels of thioredoxin transcription compared with control cells, indicating an intrinsic form of control of promoter activity by the thioredoxin system itself.
Original languageEnglish
JournalBiochemical Journal
Volume398
Issue numberPart 2
Pages (from-to)269-277
Number of pages9
ISSN0264-6021
Publication statusPublished - 2006
Externally publishedYes

Keywords

  • antioxidant responsive element
  • promoter structure
  • response rate
  • transcriptional activation
  • Primates Mammalia Vertebrata Chordata Animalia (Animals, Chordates, Humans, Mammals, Primates, Vertebrates) - Hominidae [86215] human common
  • gene
  • oxidative stress responsive element
  • specifity protein 1 binding site
  • tert-butylhydroquinone 1948-33-0
  • thioredoxin
  • 10060, Biochemistry studies - General
  • 10064, Biochemistry studies - Proteins, peptides and amino acids
  • Biochemistry and Molecular Biophysics

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