Functional identification of an intronic promoter of the human glucose-dependent insulinotropic polypeptide gene

Gene. 2010 Sep 1;463(1-2):29-40. doi: 10.1016/j.gene.2010.04.017. Epub 2010 May 7.

Abstract

Glucose-dependent insulinotropic polypeptide (GIP), a physiological incretin and enterogastrone, plays a vital role in regulating glucose-dependent insulin release from the pancreas and gastric acid secretion from the stomach. By using a transgenic mouse approach, we previously reported that the distal 1.2kb promoter region of the human GIP (hGIP) gene (-2545/-346, relative to the ATG) was able to target the transgene expression in the stomach but not in the small intestine where the majority of GIP-producing cells are located. In the present study, in order to identify the cis-acting element(s) that is/are required for intestinal expression, a 1.6kb (-1580/-) DNA fragment within the first intron of the hGIP gene was isolated and characterized in three GIP-expressing cell lines including HuTu80 (duodenal cells), PANC-1 (pancreatic ductal cells) and Hs746T (stomach cells). By 5' and 3' deletion analysis, a proximal promoter element was confined within the nucleotides -102/-1. This promoter element, functions in an orientation-dependent manner, was able to drive 15.1 and 18.3 fold increases in promoter activities in HuTu80 and PANC-1 cells, respectively. Site-directed mutation analysis indicated that the region -54/-23 was essential for promoter function while the region -22/-1 might possess opposite effects in HuTu80 and PANC-1 cells. In competitive and antibody supershift assays, interactions of the progesterone receptor (PR) and some unknown protein factors from HuTu80 and PANC-1 with the motif(s) at -54/-23 were evident. Consistent with this finding, we demonstrated the transcriptional regulation of the hGIP promoter by progesterone via the PR-B isoform and that progesterone treatment in both HuTu80 and PANC-1 cells resulted in an increase in hGIP transcript level. In addition, a sequence motif (ACATGT) residing -48/-43 was found to be responsible for the binding of potential TFII regulator(s). Taken together, our results suggest that the proximal intronic sequences contain essential cis-acting elements for the cell-specific expression of the hGIP gene.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Line, Tumor
  • Codon, Initiator
  • Duodenal Neoplasms / metabolism
  • Gastric Inhibitory Polypeptide / genetics*
  • Gastric Inhibitory Polypeptide / metabolism
  • Gene Expression Regulation
  • Humans
  • Introns
  • Mutagenesis, Site-Directed
  • Pancreatic Neoplasms / metabolism
  • Promoter Regions, Genetic*
  • Receptors, Progesterone / metabolism
  • Stomach Neoplasms / metabolism
  • Transcription Factors, TFII / metabolism

Substances

  • Codon, Initiator
  • Receptors, Progesterone
  • Transcription Factors, TFII
  • progesterone receptor B
  • Gastric Inhibitory Polypeptide