Hedgehog signaling regulation of homeodomain protein islet duodenum homeobox-1 expression in pancreatic beta-cells

Endocrinology. 2001 Mar;142(3):1033-40. doi: 10.1210/endo.142.3.8007.

Abstract

Insulin gene expression in pancreatic beta-cells is regulated by signals from developmental morphogen proteins known as hedgehogs (Hhs). By analyzing 5'-deletion insulin promoter-reporter constructs in transient transfections of clonal INS-1 beta-cells, we located activating Hh-responsive regions within the rat insulin I promoter that include the glucose-response elements Far (E2) and Flat (A2/A3). Activation of Hh signaling in INS-1 cells by ectopic Hh expression increased (and inhibition of Hh signaling with the Hh-specific inhibitor cyclopamine decreased) transcriptional activation of a multimerized FarFlat enhancer-reporter construct. In DNA-binding studies, nuclear extracts from INS-1 cells activated by ectopic Hh expression increased (and extracts from INS-1 cells treated with cyclopamine decreased) protein binding to a radiolabeled FarFlat oligonucleotide probe. An antiserum directed against the transcription factor islet duodenum homeobox-1 (IDX-1), a regulator of pancreas development and activator of the insulin gene promoter, attenuated the binding activity of Hh-responsive protein complexes. Nuclear IDX-1 protein levels on Western blots were increased by ectopic Hh expression, thereby providing a mechanism for Hh-mediated regulation of the insulin promoter. Addition of cyclopamine to INS-1 cells decreased IDX-1 messenger RNA expression. In transient transfections of a -4.5-kb mouse IDX-1 promoter-reporter construct, ectopic Hh expression increased (and cyclopamine administration decreased) transcriptional activation of the IDX-1 promoter in a dose-dependent manner. Thus, the IDX-1 gene is a direct regulatory target of Hh signaling in insulin-producing pancreatic beta-cells. We propose that Hh signaling activates the insulin gene promoter indirectly via the direct activation of IDX-1 expression. Because IDX-1 gene expression is essential for insulin gene expression, pancreatic beta-cell development, and normal glucose homeostasis, our findings that Hh signaling regulates IDX-1 expression in the endocrine pancreas suggest possible novel therapeutic approaches for diabetes mellitus.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Cell Line
  • DNA / metabolism
  • Glucose / physiology
  • Hedgehog Proteins
  • Homeodomain Proteins*
  • Insulin / genetics
  • Islets of Langerhans / metabolism*
  • Promoter Regions, Genetic / physiology
  • Proteins / physiology*
  • Rats
  • Response Elements / physiology
  • Signal Transduction / physiology*
  • Trans-Activators / genetics
  • Trans-Activators / metabolism*

Substances

  • Hedgehog Proteins
  • Homeodomain Proteins
  • Insulin
  • Proteins
  • Trans-Activators
  • pancreatic and duodenal homeobox 1 protein
  • DNA
  • Glucose