Goblet-cell-specific transcription of mouse intestinal trefoil factor gene results from collaboration of complex series of positive and negative regulatory elements

Hiroshi Itoh, Nagamu Inoue, Daniel K. Podolsky

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

Intestinal trefoil factor (ITF) is expressed selectively in intestinal goblet cells. Previous studies of the rat ITF gene identified one cis-regulatory element, designated the goblet-cell-response element (GCRE), present in the proximal region of the promoter. To identify additional cis-regulatory elements responsible for goblet-cell-specific expression, a DNA fragment containing 6353 bp of the 5'-flanking region of the mouse ITF gene was cloned and its promoter activity was examined extensively. In human and murine intestinal-derived cell lines (LS174T and CMT-93), the luciferase activities of a 6.3-kb construct were 5- and 2-fold greater than the smaller 1.8-kb construct, respectively. In contrast, the activity in non-intestinal cell lines (HepG2 and HeLa) was 2-4-fold lower than the smaller construct. In the region downstream from the 1.8-kb position, strong luciferase activities in LS174T and HepG2 cells were observed using a 201-bp construct. Interestingly, increased activity was almost completely suppressed in cells transfected with a 391-bp construct. Detailed analyses of this region revealed the existence of a 11-bp positive regulatory element (-181 to -170; ACCTCTTCCTG) and a 9-bp negative regulatory element (-208 to -200; ATTGACAGA) in addition to the GCRE. All three elements were well conserved among human, rat and mouse ITF gene promoters. In addition, a mutant 1.8-kb construct in which the negative regulatory region was deleted yielded the same approximate luciferase activity as a 6.3-kb construct, suggesting binding of a goblet-cell-specific silencer inhibitor (SI) between -6.3 and -1.8 kb. The SI present in goblet cells may block the silencers' binding to the pre-initiation complex and allow increased transcriptional activity driven by specific and non-specific enhancers. High-level expression of the mouse ITF gene specifically in intestinal goblet cells may be achieved through the combined effects of these regulatory elements.

Original languageEnglish
Pages (from-to)461-472
Number of pages12
JournalBiochemical Journal
Volume341
Issue number2
DOIs
Publication statusPublished - 1999 Jul 15
Externally publishedYes

Fingerprint

Goblet Cells
Transcription
Genes
Luciferases
Response Elements
Cells
5' Flanking Region
Nucleic Acid Regulatory Sequences
Genetic Promoter Regions
Rats
Cell Line
Hep G2 Cells
Trefoil Factor-3
DNA

Keywords

  • Colon
  • Enhancer
  • Epithelium
  • Promoter
  • Silencer

ASJC Scopus subject areas

  • Biochemistry

Cite this

Goblet-cell-specific transcription of mouse intestinal trefoil factor gene results from collaboration of complex series of positive and negative regulatory elements. / Itoh, Hiroshi; Inoue, Nagamu; Podolsky, Daniel K.

In: Biochemical Journal, Vol. 341, No. 2, 15.07.1999, p. 461-472.

Research output: Contribution to journalArticle

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