Nature Genetics 38, 312 - 319 (2006)
Published online: 12 February 2006; | doi:10.1038/ng1745
Feedback repression is required for mammalian circadian clock functionTrey K Sato1, 2, 3, 9, Rikuhiro G Yamada4, 9, Hideki Ukai4, 9, Julie E Baggs5, Loren J Miraglia1, Tetsuya J Kobayashi4, David K Welsh2, 6, 7, Steve A Kay2, 5, Hiroki R Ueda4
& John B Hogenesch1, 3, 5, 81
Genomics Institute of the Novartis Research Foundation, 10675 John J. Hopkins Dr., San Diego, California 92121, USA. 2
Department of Cell Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA. 3
Department of Genome Technology, The Scripps Research Institute, 5353 Parkside Drive, RF1, Jupiter, Florida 33458, USA. 4
Laboratory for Systems Biology, Center for Developmental Biology, RIKEN, 2-2-3 Minatojima-minamimachi, Chuo-ku, Kobe, Hyogo, 650-0047, Japan. 5
Department of Biochemistry, The Scripps Research Institute, 5353 Parkside Drive, RF1, Jupiter, Florida 33458, USA. 6
Department of Psychiatry, University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92037, USA. 7
The Veterans Administration San Diego Healthcare System, 3350 La Jolla Village Drive, San Diego, California 92161, USA. 8
Department of Neuropharmacology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA. 9
These authors contributed equally to this work.
Correspondence should be addressed to Hiroki R Ueda uedah-tky@umin.ac.jp or John B Hogenesch hogenesch@scripps.edu Direct evidence for the requirement of transcriptional feedback repression in circadian clock function has been elusive. Here, we developed a molecular genetic screen in mammalian cells to identify mutants of the circadian transcriptional activators CLOCK and BMAL1, which were uncoupled from CRYPTOCHROME (CRY)-mediated transcriptional repression. Notably, mutations in the PER-ARNT-SIM domain of CLOCK and the C terminus of BMAL1 resulted in synergistic insensitivity through reduced physical interactions with CRY. Coexpression of these mutant proteins in cultured fibroblasts caused arrhythmic phenotypes in population and single-cell assays. These data demonstrate that CRY-mediated repression of the CLOCK/BMAL1 complex activity is required for maintenance of circadian rhythmicity and provide formal proof that transcriptional feedback is required for mammalian clock function.
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