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  • The EMBO Journal (2002) 21, 1301 - 1314
  • doi:10.1093/emboj/21.6.1301

Nucleocytoplasmic shuttling and mCRY-dependent inhibition of ubiquitylation of the mPER2 clock protein

Kazuhiro Yagita1, Filippo Tamanini2, Maya Yasuda1, Jan H. J. Hoeijmakers2, Gijsbertus T. J. van der Horst2 and Hitoshi Okamura1

  1. Division of Molecular Brain Science, Department of Brain Sciences, Kobe University Graduate School of Medicine, Kobe 650-0017, Japan
  2. MGC, Department of Cell Biology and Genetics, Erasmus University, PO Box 1738, 3000 DR Rotterdam, The Netherlands

Correspondence to:

Hitoshi Okamura, E-mail: okamurah@kobe-u.ac.jp

Gijsbertus T. J. van der Horst, E-mail: vanderhorst@gen.fgg.eur.nl

Received 27 November 2001; Accepted 18 January 2002; Revised 28 December 2001

The core oscillator generating circadian rhythms in eukaryotes is composed of transcription–translation-based autoregulatory feedback loops in which clock gene products negatively affect their own expression. A key step in this mechanism involves the periodic nuclear accumulation of clock proteins following their mRNA rhythms after approx6 h delay. Nuclear accumulation of mPER2 is promoted by mCRY proteins. Here, using COS7 cells and mCry1/mCry2 double mutant mouse embryonic fibroblasts transiently expressing GFP-tagged (mutant) mPER2, we show that the protein shuttles between nucleus and cytoplasm using functional nuclear localization and nuclear export sequences. Moreover, we provide evidence that mCRY proteins prevent ubiquitylation of mPER2 and subsequent degradation of the latter protein by the proteasome system. Interestingly, mPER2 in turn prevents ubiquitylation and degradation of mCRY proteins. On the basis of these data we propose a model in which shuttling mPER2 is ubiquitylated and degraded by the proteasome unless it is retained in the nucleus by mCRY proteins.

  • Keywords:

    • circadian clock,
    • cryptochrome,
    • nuclear–cytoplasmic shuttling,
    • period,
    • ubiquitylation