Identification of negative transcriptional factor E4BP4-binding site in the mouse circadian-regulated gene Mdr2

• Published in: Neuroscience research 2008-03, Vol.60 (3), p.307-313
• Main Authors: Kotaka, Maki, Onishi, Yoshiaki, Ohno, Tomoya, Akaike, Toshihiro, Ishida, Norio
• Format: Article
• Language: English
• Subjects: Animals; ATP Binding Cassette Transporter, Subfamily B - genetics; ATP-binding cassette (ABC) transporter; ATP-Binding Cassette Sub-Family B Member 4; Basic-Leucine Zipper Transcription Factors - metabolism; Carcinoma, Hepatocellular; Cell Line, Tumor; Circadian; Circadian Rhythm - genetics; cis-Element; CLOCK Proteins; E4BP4; Humans; Liver - physiology; Liver Neoplasms; Male; Mice; Mice, Inbred C57BL; Mice, Inbred ICR; Mice, Mutant Strains; Multidrug resistance 2 (Mdr2); Negative transcription; Promoter Regions, Genetic - physiology; RNA, Messenger - metabolism; Trans-Activators - genetics; Transcriptional Activation - physiology
• ISSN: 0168-0102; 1872-8111
• DOI: 10.1016/j.neures.2007.11.014

The hepatic transporter Mdr2 is an ATP-binding cassette transporter which excretes phosphatidylcholine into the bile. We showed that the level of Mdr2 mRNA oscillated in circadian fashion in mouse liver whereas such oscillation was dampened in the liver of Clock mutants. To examine transcriptional regulation of the Mdr2 gene we performed luciferase reporter assays using plasmid constructs containing the 5′-flanking region of the Mdr2 gene. Reporter assays using deletion constructs demonstrated that E4BP4 represses the transcriptional activity of the promoter including the D1 and D2 sites within four putative E4BP4-binding sites. Chromatin immunoprecipitation and gel shift assays showed that E4BP4 binds to the D2 site, but not to the D1 site. These data suggested that E4BP4 is a negative transcription factor for circadian Mdr2 mRNA expression.