Expression of the Robo4 receptor in endothelial cells is regulated by two AP-1 protein complexes

Biochem Biophys Res Commun. 2015 Nov 27;467(4):987-91. doi: 10.1016/j.bbrc.2015.10.029. Epub 2015 Oct 14.


Roundabout4 (Robo4) is an endothelial cell-specific gene that plays an important role in endothelial cell stability. We previously identified a 3-kb Robo4 promoter and demonstrated the importance of its proximal region in regulating Robo4 gene expression. To investigate the role of the upstream promoter in Robo4 gene regulation, we searched evolutionarily conserved promoter regions by phylogenetic footprinting and identified three conserved promoter regions. The most upstream region included a conserved AP-1 binding motif at position -2875. A mutation in the AP-1 motif significantly decreased Robo4 promoter activity in a transient reporter assay. An electrophoretic mobility shift assay and a chromatin immunoprecipitation assay demonstrated binding of a c-Jun/c-Jun complex and a c-Jun/Fra-1 complex to the AP-1 motif. Knockdown experiments using siRNA revealed that both c-Jun/c-Jun and c-Jun/Fra-1 complexes regulate Robo4 gene expression, and that the c-Jun/c-Jun complex is essential for maximum promoter activation. Collectively, these results indicate that AP-1 complexes regulate Robo4 gene expression in endothelial cells.

Keywords: AP-1; Endothelial cell; Fra-1; Robo4; c-Jun.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Cells, Cultured
  • Endothelium, Vascular / cytology
  • Endothelium, Vascular / metabolism*
  • Gene Expression Regulation / physiology
  • Humans
  • Promoter Regions, Genetic
  • Receptors, Cell Surface / genetics
  • Receptors, Cell Surface / metabolism*
  • Sequence Homology, Nucleic Acid
  • Transcription Factor AP-1 / metabolism
  • Transcription Factor AP-1 / physiology*


  • ROBO4 protein, human
  • Receptors, Cell Surface
  • Transcription Factor AP-1