Analysis and synthesis of high-amplitude Cis-elements in the mammalian circadian clock

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 105; no. 39; pp. 14946 - 14951
• Main Authors: Kumaki, Yuichi, Ukai-Tadenuma, Maki, Uno, Ken-ichiro D, Nishio, Junko, Masumoto, Koh-hei, Nagano, Mamoru, Komori, Takashi, Shigeyoshi, Yasufumi, Hogenesch, John B, Ueda, Hiroki R
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 30.09.2008; National Acad Sciences
• Subjects: Animals; ARNTL Transcription Factors; Base Sequence; Basic Helix-Loop-Helix Transcription Factors - metabolism; Basic-Leucine Zipper Transcription Factors; Binding sites; Binding, Competitive; Biochemistry; Biological Sciences; Bioluminescence; Circadian Rhythm - genetics; CLOCK Proteins; Comparative genomics; Databases, Genetic; DNA-Binding Proteins - metabolism; Enhancer Elements, Genetic; Evolution; Gene expression; Gene Expression Regulation; Genes; Genomes; Genomics; Humans; Mammals; Mice; Promoter Regions, Genetic; Response elements; Sequence Analysis, DNA; Trans-Activators - metabolism; Transcription factors; Transcription Factors - metabolism
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0802636105

Mammalian circadian clocks consist of regulatory loops mediated by Clock/Bmal1-binding elements, DBP/E4BP4 binding elements, and RevErbA/ROR binding elements. As a step toward system-level understanding of the dynamic transcriptional regulation of the oscillator, we constructed and used a mammalian promoter/enhancer database (http://promoter.cdb.riken.jp/) with computational models of the Clock/Bmal1-binding elements, DBP/E4BP4 binding elements, and RevErbA/ROR binding elements to predict new targets of the clock and subsequently validated these targets at the level of the cell and organism. We further demonstrated the predictive nature of these models by generating and testing synthetic regulatory elements that do not occur in nature and showed that these elements produced high-amplitude circadian gene regulation. Biochemical experiments to characterize these synthetic elements revealed the importance of the affinity balance between transactivators and transrepressors in generating high-amplitude circadian transcriptional output. These results highlight the power of comparative genomics approaches for system-level identification and knowledge-based design of dynamic regulatory circuits.