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Role for Slimb in the degradation of Drosophila Period protein phosphorylated by Doubletime

Nature volume 420, pages 673–678 (2002)Cite this article

Abstract

Protein phosphorylation has a key role in modulating the stabilities of circadian clock proteins in a manner specific to the time of day1. A conserved feature of animal clocks is that Period (Per) proteins undergo daily rhythms in phosphorylation and levels2,3, events that are crucial for normal clock progression4,5,6,7. Casein kinase Iε (CKIε) has a prominent role in regulating the phosphorylation and abundance of Per proteins in animals8. This was first shown in Drosophila with the characterization of Doubletime (Dbt), a homologue of vertebrate casein kinase Iε4,6. However, it is not clear how Dbt regulates the levels of Per. Here we show, using a cell culture system, that Dbt promotes the progressive phosphorylation of Per, leading to the rapid degradation of hyperphosphorylated isoforms by the ubiquitin–proteasome pathway. Slimb, an F-box/WD40-repeat protein functioning in the ubiquitin–proteasome pathway9,10 interacts preferentially with phosphorylated Per and stimulates its degradation. Overexpression of slimb or expression in clock cells of a dominant-negative version of slimb disrupts normal rhythmic activity in flies. Our findings suggest that hyperphosphorylated Per is targeted to the proteasome by interactions with Slimb.

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Figure 1: Progressive phosphorylation of Per by Dbt in cultured Drosophila cells.
Figure 2: Alterations in Per phosphorylation and degradation by Tim and mutant forms of Dbt.
Figure 3: Slimb interacts preferentially with Dbt-phosphorylated Per and mediates its rapid degradation.
Figure 4: Accumulation of DBT-dependent highly phosphorylated Per by silencing endogenous slimb activity.
Figure 5: Altered locomotor activity rhythms in tub-slimb flies.
Figure 6: Altered locomotor activity rhythms in tim-gal4 × (UAS)Slimb*Fbox flies.

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Acknowledgements

We thank S. Kay for the pAct-per and pAct-tim constructs, A. Seghal for the pCasper-hs-Ub/HA construct, J. Hall for the tim-gal4-27 and tim-gal4-62 lines, and M. Young for the tim-(UAS)-gal4 line. The work was supported by the Searle Scholar Program and the UTSW Endowed Scholar Program to J.J., and by a grant from the National Institutes of Health to I.E.

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Authors and Affiliations

  1. Graduate Program in Physiology and Neurobiology, Rutgers University, Center for Advanced Biotechnology and Medicine, 679 Hoes Lane, Piscataway, New Jersey, 08854, USA

    Hyuk Wan Ko

  2. Department of Molecular Biology and Biochemistry, Rutgers University, Center for Advanced Biotechnology and Medicine, 679 Hoes Lane, Piscataway, New Jersey, 08854, USA

    Isaac Edery

  3. Center for Developmental Biology and Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, Texas, 75390, USA

    Jin Jiang

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Correspondence to Isaac Edery.

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Ko, H., Jiang, J. & Edery, I. Role for Slimb in the degradation of Drosophila Period protein phosphorylated by Doubletime. Nature 420, 673–678 (2002). https://doi.org/10.1038/nature01272

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