title: Nucleocytoplasmic shuttling and mCRY-dependent inhibition of ubiquitylation of the mPER2 clock protein.
abstract: 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 approximately 6 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.
authors.affiliation: Division of Molecular Brain Science, Department of Brain Sciences, Kobe University Graduate School of Medicine, Kobe 650-0017, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Yagita
authors.forename: Filippo
authors.initials: F
authors.lastname: Tamanini
authors.forename: Maya
authors.initials: M
authors.lastname: Yasuda
authors.forename: Jan H J
authors.initials: JH
authors.lastname: Hoeijmakers
authors.forename: Gijsbertus T J
authors.initials: GT
authors.lastname: van der Horst
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Okamura
mesh_terms: D016475:3T3 Cells; D021581:Active Transport, Cell Nucleus; D000818:Animals; D001683:Biological Clocks; D019556:COS Cells; D018797:Cell Cycle Proteins; D002467:Cell Nucleus / Q000378:metabolism; D002522:Chlorocebus aethiops; D056931:Cryptochromes; D003593:Cytoplasm / Q000378:metabolism; D029721:Drosophila Proteins; D005136:Eye Proteins; D005420:Flavoproteins / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D029543:Nucleocytoplasmic Transport Proteins / Q000235:genetics / Q000378:metabolism*; D056950:Period Circadian Proteins; D017956:Photoreceptor Cells, Invertebrate; D021382:Protein Sorting Signals; D043562:Receptors, G-Protein-Coupled; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors; D025801:Ubiquitin / Q000378:metabolism*
chemical_list: D018797:Cell Cycle Proteins; D056931:Cryptochromes; D029721:Drosophila Proteins; D005136:Eye Proteins; D005420:Flavoproteins; D009687:Nuclear Proteins; D029543:Nucleocytoplasmic Transport Proteins; C421902:PER2 protein, human; C513312:Per2 protein, mouse; D056950:Period Circadian Proteins; D021382:Protein Sorting Signals; D043562:Receptors, G-Protein-Coupled; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D025801:Ubiquitin; C119828:cry protein, Drosophila