Role of chromatin accessibility in the occupancy and transcription of the insulin gene by the pancreatic and duodenal homeobox factor 1

Joshua Francis, Daniella A. Babu, Tye G. Deering, Swarup K. Chakrabarti, James C. Garmey, Carmella Evans-Molina, David G. Taylor, Raghavendra G. Mirmira

Research output: Contribution to journalArticlepeer-review

13 Scopus citations

Abstract

The pancreatic and duodenal homeobox factor 1 (Pdx-1) is a Hox-like transcription factor that is responsible for the activation of the insulin gene. Previous studies have demonstrated the interaction in vitro of Pdx-1 with short (20-40 nucleotide) DNA fragments corresponding to A boxes of the insulin promoter. Precisely how Pdx-1 binds to DNA in the complex milieu of chromatin, however, has never been studied. In this study, we explored how Pdx-1-DNA interactions might be influenced by chromatin accessibility at the insulin gene in β-cells (βTC3) vs. pancreatic ductal cells (mPAC). We demonstrate that Pdx-1 occupies the endogenous insulin promoter in βTC3 cells but not in mPAC cells, a finding that is independent of the intracellular Pdx-1 protein concentration. Based on micrococcal nuclease protection assays, the difference in promoter binding between the two cell types appears to be secondary to chromatin accessibility at predicted Pdx-1 binding sites between bp -126 to -296 (relative to the transcriptional start site) of the insulin promoter. Binding studies using purified Pdx-1 and reconstituted chromatin in vitro suggest that the positioning of a nucleosome(s) within this crucial region of the promoter might account for differences in chromatin accessibility. Consistent with these observations, fluorescence colocalization studies show that Pdx-1 does not occupy regions of compacted, nucleosome-rich chromatin within the nucleus. Our findings suggest a model whereby insulin transcription in the β-cell is at least partially facilitated by enhanced chromatin accessibility within a crucial regulatory region between bp -126 to -296, thereby permitting occupancy by transactivators such as Pdx-1.

Original languageEnglish (US)
Pages (from-to)3133-3145
Number of pages13
JournalMolecular Endocrinology
Volume20
Issue number12
DOIs
StatePublished - Dec 2006

ASJC Scopus subject areas

  • Molecular Biology
  • Endocrinology

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