Abstract
Protein phosphorylation has a key role in modulating the stabilities of circadian clock proteins in a manner specific to the time of day. A conserved feature of animal clocks is that Period (Per) proteins undergo daily rhythms in phosphorylation and levels, events that are crucial for normal clock progression. Casein kinase Iepsilon (CKIepsilon) has a prominent role in regulating the phosphorylation and abundance of Per proteins in animals. This was first shown in Drosophila with the characterization of Doubletime (Dbt), a homologue of vertebrate casein kinase Iepsilon. 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 pathway 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.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Animals, Genetically Modified
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Biological Clocks
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Casein Kinase 1 epsilon*
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / metabolism*
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Cell Line
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Circadian Rhythm
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Cysteine Endopeptidases / metabolism
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Drosophila Proteins / genetics
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Drosophila Proteins / metabolism*
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Drosophila melanogaster / cytology
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Drosophila melanogaster / genetics
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Drosophila melanogaster / metabolism*
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Drosophila melanogaster / physiology
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Insect Proteins / genetics
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Insect Proteins / metabolism*
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Motor Activity
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Multienzyme Complexes / metabolism
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Mutation / genetics
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Nuclear Proteins / chemistry
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Nuclear Proteins / metabolism*
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Period Circadian Proteins
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Phosphorylation
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Proteasome Endopeptidase Complex
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Protein Binding
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Protein Isoforms / metabolism
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Protein Kinases / genetics
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Protein Kinases / metabolism*
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RNA Interference
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Substrate Specificity
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Ubiquitin-Protein Ligases*
Substances
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Cell Cycle Proteins
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Drosophila Proteins
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Insect Proteins
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Multienzyme Complexes
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Nuclear Proteins
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PER protein, Drosophila
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Period Circadian Proteins
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Protein Isoforms
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dco protein, Drosophila
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slmb protein, Drosophila
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tim protein, Drosophila
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Ubiquitin-Protein Ligases
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Protein Kinases
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Casein Kinase 1 epsilon
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Cysteine Endopeptidases
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Proteasome Endopeptidase Complex