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KMID : 0613820160260010042
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Journal of Life Science
2016 Volume.26 No. 1 p.42 ~ p.49
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Molecular Cloning and Characterization of Bovine CYP26A1 Promoter
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Kwak In-Seok
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Abstract
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The retinoic acid (RA) plays an important role in the growth and development of many cells, and bioactive RA concentration is regulated by several enzymes, including CYP26A1. The expression of the CYP26A1 gene is regulated by RA, and the CYP26A1 gene is one of the candidates for RA-responsive genes. Although CYP26A1 genes are cloned from several animals, cloning of the CYP26A1 gene from cows has not been reported yet. The promoter region of CYP26A1 from cows was cloned by PCR and analyzed by sequence alignment with human and mouse CYP26A1. The RA-responsive element (RARE), DR-5 (TGAACTttgggTGAACT), was located in this region and was perfectly conserved. The promoter region of bovine CYP26A1, which contains DR-5, was ligated to the luciferase reporter gene on transient transfection assays. The expression of CYP26A1-Luc promoter was activated by ATRA treatment in lung-derived mtCC cells. Co-transfection with RAR-α or -β with ATRA significantly activates the expression of CYP26A1-Luc promoter; however, it was less effective with either RAR-γ or RXR-γ. In addition, the endogenous gene expressions measured by Q-RT-PCR in mtCC cells were not significantly affected by ATRA treatment for 2 days; however, the expression of the endogenous CYP26A1 gene was diminished sharply at day 3 with ATRA treatment. In conclusion, the promoter region of bovine CYP26A1 contains conserved DR-5 RARE, which functions as a binding site for RAR-α or ?β, and it is involved in the regulation of CYP26A1 gene expression and the control of RA signaling in mtCC cells.
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KEYWORD
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ATRA, CYP26A1, RAR, RARE (responsive element), RXR
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