Molecules and Cells

Korean Society for Molecular and Cellular Biology (한국분자세포생물학회)
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Identification of an Enhancer Critical for the ephirn-A5 Gene Expression in the Posterior Region of the Mesencephalon

  • Park, Eunjeong (Department of Biological Sciences, Sookmyung Women's University) ;
  • Noh, Hyuna (Department of Biological Sciences, Sookmyung Women's University) ;
  • Park, Soochul (Department of Biological Sciences, Sookmyung Women's University)
  • Received : 2017.03.29
  • Accepted : 2017.05.11
  • Published : 2017.06.30
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Abstract

Ephrin-A5 has been implicated in the regulation of brain morphogenesis and axon pathfinding. In this study, we used bacterial homologous recombination to express a LacZ reporter in various ephrin-A5 BAC clones to identify elements that regulate ephrin-A5 gene expression during mesencephalon development. We found that there is mesencephalon-specific enhancer activity localized to a specific +25.0 kb to +30.5 kb genomic region in the first intron of ephrin-A5. Further comparative genomic analysis indicated that two evolutionary conserved regions, ECR1 and ECR2, were present within this 5.5 kb region. Deletion of ECR1 from the enhancer resulted in disrupted mesencephalon-specific enhancer activity in transgenic embryos. We also found a consensus binding site for basic helix-loop-helix (bHLH) transcription factors (TFs) in a highly conserved region at the 3'-end of ECR1. We further demonstrated that specific deletion of the bHLH TF binding site abrogated the mesencephalon-specific enhancer activity in transgenic embryos. Finally, both electrophoretic mobility shift assay and luciferase-based transactivation assay revealed that the transcription factor Ascl1 bound the bHLH consensus binding site in the mesencephalon-specific ephrin-A5 enhancer in vitro. Together, these results suggest that the bHLH TF binding site in ECR1 is involved in the positive regulation of ephrin-A5 gene expression during the development of the mesencephalon.

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References

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