International Mammalian Genome Society

The 14th International Mouse Genome Conference (2000)


B21. Molecular Mechanisms of Positive and Negative Regulation of Per1 Transcription

Akiko Hida-Fukuda1,2, Nobuya Koike2, Matsumi Hirose1,2, Masahira Hattori1,2, Yoshiyuki Sakaki1,2, and Hajime Tei2
1Human Genome Research Group, Genomic Sciences Center, RIKEN, 1-15-1 Kitasato, Sagamihara-shi, Kanagawa 228-8555, Japan;
2
Laboratory of Functional Genomics, Human Genome Center, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan

Circadian clocks control the daily oscillations of biochemical, physiological and behavioral rhythms. These rhythms are generated by endogenous pacemakers and are synchronized by light-dark cycles. The transcription of mouse Per1 (mPer1) in the suprachiasmatic nucleus (SCN) oscillates autonomously and is induced immediately under light exposure. The results indicate that the daily oscillation of Per1 plays an important role in the maintenance of circadian rhythms. Per1 transcription is induced by the CLOCK-BMAL1 complex and is repressed by CRY1 and CRY2. To elucidate the molecular mechanisms underlying Per1 regulation, the mPer1 promoter was analyzed by a transient transfection assay with mouse NIH3T3 cells using mPer1: luciferase (luc) reporter genes. Both of the human and mouse Per1 genes (hPER1 and mPer1) consisted of 23 exons spanning approximately 16 kb. Their structures showed strong similarity to each other and six conserved segments containing five E-boxes (the binding site for the CLOCK-BMAL1 complex) were identified in the human and mouse Per1 promoter regions. Introduction of deletions or point mutations into the five E-boxes showed that each of the five E-boxes was functional for the mPer1 induction by CLOCK and BMAL1, and gel shift analysis demonstrated that the CLOCK-BMAL1 complex bound to the E-box. Transient transfection assays using the deletion mutants showed that both CRY1 and CRY2 completely repressed the CLOCK-BMAL1-induced expression independent of the mPer1 promoter region. In addition, the binding of CRY1 and CRY2 to the E-box was not detected in gel shift analysis. These results clearly indicate that mPer1 transcription is induced by the CLOCK-BMAL1 complex through directly binding each of the five E-boxes, and is repressed not by the interaction of any cis-elements in the mPer1 promoter with CRY1 and CRY2.

 


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