VEGF/Flk1 signaling cascade transactivates Etv2 gene expression

PLoS One. 2012;7(11):e50103. doi: 10.1371/journal.pone.0050103. Epub 2012 Nov 19.

Abstract

Previous reports regarding the genetic hierarchy between Ets related protein 71 (Er71/Etv2) and Flk1 is unclear. In the present study, we pursued a genetic approach to define the molecular cascade between Etv2 and Flk1. Using a transgenic Etv2-EYFP reporter mouse, we examined the expression pattern of Etv2 relative to Flk1 in the early conceptus. Etv2-EYFP was expressed in subset of Flk1 positive cells during primitive streak stages, suggesting that Flk1 is upstream of Etv2 during gastrulation. Analysis of reporter gene expression in Flk1 and Etv2 mutant mice further supports the hypothesis that Flk1 is necessary for Etv2 expression. The frequency of cells expressing Flk1 in Etv2 mutants is only modestly altered (21% decrease), whereas expression of the Etv2-EYFP transgenic reporter was severely reduced in the Flk1 null background. We further demonstrate using transcriptional assays that, in the presence of Flk1, the Etv2 promoter is activated by VEGF, the Flk1 ligand. Pharmacological inhibition studies demonstrate that VEGF mediated activation is dependent on p38 MAPK, which activates Creb. We identify the VEGF response element in the Etv2 promoter and demonstrate that Creb binds to this motif by EMSA and ChIP assays. In summary, we provide new evidence that VEGF activates Etv2 by signaling through Flk1, which activates Creb through the p38 MAPK signaling cascade.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Cyclic AMP Response Element-Binding Protein / genetics*
  • Cyclic AMP Response Element-Binding Protein / metabolism
  • Embryo, Mammalian
  • Gastrulation / genetics*
  • Gene Expression Regulation, Developmental
  • Mice
  • Mice, Transgenic
  • Molecular Sequence Data
  • Protein Binding
  • Response Elements
  • Signal Transduction
  • Transcription Factors / genetics*
  • Transcription Factors / metabolism
  • Transcriptional Activation
  • Vascular Endothelial Growth Factor A / genetics*
  • Vascular Endothelial Growth Factor A / metabolism
  • Vascular Endothelial Growth Factor Receptor-2 / genetics*
  • Vascular Endothelial Growth Factor Receptor-2 / metabolism
  • p38 Mitogen-Activated Protein Kinases / genetics*
  • p38 Mitogen-Activated Protein Kinases / metabolism

Substances

  • Creb1 protein, mouse
  • Cyclic AMP Response Element-Binding Protein
  • ER71 protein, mouse
  • Transcription Factors
  • Vascular Endothelial Growth Factor A
  • vascular endothelial growth factor A, mouse
  • Vascular Endothelial Growth Factor Receptor-2
  • p38 Mitogen-Activated Protein Kinases