Relative contribution of PDX-1, MafA and E47/b2 to the regulation of the human insulin promoter.

Hilary Docherty, Colin Hay, Angela Louise Ferguson, John Barrow, Elaine Durward, Kevin Docherty

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64 Citations (Scopus)

Abstract

The insulin promoter binds a number of tissue-specific and ubiquitous transcription factors. Of these, the homoeodomain protein PDX-1 (pancreatic duodenal homeobox factor-1), the basic leucine zipper protein MafA and the basic helix-loop-helix heterodimer E47/BETA2 (beta-cell E box transactivator 2; referred to here as beta 2) bind to important regulatory sites. Previous studies have shown that PDX-1 can interact synergistically with E47 and beta 2 to activate the rat insulin I promoter. The aim of the present study was to determine the relative contribution of PDX-1, MafA and E47/beta 2 in regulating the human insulin promoter, and whether these factors could interact synergistically in the context of the human promoter. Mutagenesis of the PDX-1, MafA and E47/beta 2 binding sites reduced promoter activity by 60, 74 and 94% respectively, in INS-1 beta-cells. In the islet glucagonoma cell line alpha TC1.6, overexpression of PDX-1 and MafA separately increased promoter activity approx. 2.5-3-fold, and in combination approx. 6-fold, indicating that their overall effect was additive. Overexpression of E47 and beta 2 had no effect. In HeLa cells, PDX-1 stimulated the basal promoter by approx. 40-fold, whereas MafA, E47 and beta 2 each increased activity by less than 2-fold. There was no indication of any synergistic effects on the human insulin promoter. On the other hand, the rat insulin I promoter and a mutated version of the human insulin promoter, in which the relevant regulatory elements were separated by the same distances as in the rat insulin I promoter, did exhibit synergy. PDX-1 was shown further to activate the endogenous insulin I gene in alpha TC1.6 cells, whereas MafA activated the insulin 2 gene. In combination, PDX-1 and MafA activated both insulin genes. Chromatin immunoprecipitation assays confirmed that PDX-1 increased the association of acetylated histones H3 and H4 with the insulin I gene and MafA increased the association of acetylated histone H3 with the insulin 2 gene.

Original languageEnglish
Pages (from-to)813-820
Number of pages8
JournalBiochemical Journal
Volume389
Issue number3
DOIs
Publication statusPublished - 1 Aug 2005

Keywords

  • animals
  • basic Helix-Loop-Helix transcription Factors
  • cell line, tumor
  • DNA-binding proteins
  • gene expression regulation
  • HMGB proteins
  • homeodomain proteins
  • humans
  • insulin
  • Maf transcription factors, Large
  • Promoter Regions, Genetic
  • TCF transcription factors
  • trans-activators
  • transcription factors
  • diabetes mellitus
  • gemne transcription
  • islets of Langerhans
  • PDX-1

Cite this

Docherty, H., Hay, C., Ferguson, A. L., Barrow, J., Durward, E., & Docherty, K. (2005). Relative contribution of PDX-1, MafA and E47/b2 to the regulation of the human insulin promoter. Biochemical Journal, 389(3), 813-820. https://doi.org/10.1042/BJ20041891