Stimulation of B-Raf increases c-Jun and c-Fos expression and upregulates AP-1-regulated gene transcription in insulinoma cells

Mol Cell Endocrinol. 2018 Sep 5;472:126-139. doi: 10.1016/j.mce.2017.12.003. Epub 2017 Dec 8.

Abstract

Stimulation of pancreatic β-cells with glucose activates the protein kinases B-Raf and extracellular signal-regulated protein kinase that participate in glucose sensing. Inhibition of both kinases results in impairment of glucose-regulated gene transcription. To analyze the signaling pathway controlled by B-Raf, we expressed a conditionally active form of B-Raf in INS-1 insulinoma cells. Here, we show that stimulation of B-Raf strongly activated the transcription factor AP-1 which is accompanied by increased c-Jun and c-Fos promoter activities, an upregulation of c-Jun and c-Fos biosynthesis, and elevated transcriptional activation potentials of c-Jun and c-Fos. Mutational analysis identified the AP-1 sites within the c-Jun promoter and the serum response element (SRE) within the c-Fos promoter as the essential genetic elements connecting B-Raf stimulation with AP-1 activation. In line with this, the transcriptional activation potential of the SRE-binding protein Elk-1 was increased following B-Raf activation. The signal pathway from B-Raf to AP-1 required the activation of c-Jun. We identified the cyclin D1 gene as a delayed response gene for AP-1 following stimulation of B-Raf in insulinoma cells. Moreover, MAP kinase phosphatase-1 and the Ca2+/calmodulin-dependent protein phosphatase calcineurin were identified to function as shut-off-devices for the signaling cascade connecting B-Raf stimulation with the activation of AP-1. The fact that stimulation with glucose, activation of L-type voltage-gated Ca2+ channels, and stimulation of B-Raf all trigger an activation of AP-1 indicates that AP-1 is a point of convergence of signaling pathways in β-cell.

Keywords: AP-1; Calcineurin; Elk-1; MKP-1; c-Fos; c-Jun.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Calcineurin / metabolism
  • Calcium Channels / metabolism
  • Cell Line, Tumor
  • Cyclin D1 / genetics
  • Dual Specificity Phosphatase 1 / metabolism
  • Gene Expression Regulation, Neoplastic
  • Insulinoma / genetics*
  • Models, Biological
  • Mutation / genetics
  • Promoter Regions, Genetic
  • Proto-Oncogene Proteins B-raf / metabolism*
  • Proto-Oncogene Proteins c-fos / genetics*
  • Proto-Oncogene Proteins c-fos / metabolism
  • Proto-Oncogene Proteins c-jun / genetics*
  • Proto-Oncogene Proteins c-jun / metabolism
  • Rats
  • Serum Response Element / genetics
  • Transcription Factor AP-1 / genetics*
  • Transcription Factor AP-1 / metabolism
  • Transcription, Genetic*
  • Transcriptional Activation / genetics
  • Up-Regulation / genetics*
  • ets-Domain Protein Elk-1 / metabolism

Substances

  • Calcium Channels
  • Proto-Oncogene Proteins c-fos
  • Proto-Oncogene Proteins c-jun
  • Transcription Factor AP-1
  • ets-Domain Protein Elk-1
  • Cyclin D1
  • Proto-Oncogene Proteins B-raf
  • Calcineurin
  • Dual Specificity Phosphatase 1