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Protein phosphatase 4 controls circadian clock dynamics by modulating CLOCK/BMAL1 activity
Sabrina Klemz1, Thomas Wallach1, Sandra Korge1
1Laboratory of Chronobiology, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin,10117 Berlin, Germany.
Abstract:
In all organisms with circadian clocks, post-translational modifications of clock proteins control the dynamics of circadian rhythms, with phosphorylation playing a dominant role. All major clock proteins are highly phosphorylated, and many kinases have been described to be responsible. In contrast, it is largely unclear whether and to what extent their counterparts, the phosphatases, play an equally crucial role. To investigate this, we performed a systematic RNAi screen in human cells and identified protein phosphatase 4 (PPP4) with its regulatory subunit PPP4R2 as critical components of the circadian system in both mammals and Drosophila Genetic depletion of PPP4 shortens the circadian period, whereas overexpression lengthens it. PPP4 inhibits CLOCK/BMAL1 transactivation activity by binding to BMAL1 and counteracting its phosphorylation. This leads to increased CLOCK/BMAL1 DNA occupancy and decreased transcriptional activity, which counteracts the "kamikaze" properties of CLOCK/BMAL1. Through this mechanism, PPP4 contributes to the critical delay of negative feedback by retarding PER/CRY/CK1δ-mediated inhibition of CLOCK/BMAL1.
Insights
Protein phosphatase 4 (PPP4) regulates circadian rhythms by inhibiting CLOCK/BMAL1 activity. This phosphatase delays negative feedback, ensuring proper circadian clock function in mammals and Drosophila.
Area of Science:
- Chronobiology
- Molecular Biology
- Biochemistry
Background:
- Circadian rhythms are governed by post-translational modifications of clock proteins, primarily phosphorylation.
- While kinases are well-studied, the role of phosphatases in circadian regulation remains largely unknown.
Purpose of the Study:
- To investigate the role of phosphatases in circadian clock function.
- To identify specific phosphatases involved in regulating circadian rhythms.
Main Methods:
- Systematic RNA interference (RNAi) screen in human cells.
- Functional analysis of identified phosphatases in mammalian and Drosophila circadian systems.
Main Results:
- Protein phosphatase 4 (PPP4) and its regulatory subunit PPP4R2 were identified as critical circadian components.
- Depletion of PPP4 shortened the circadian period; overexpression lengthened it.
- PPP4 inhibits CLOCK/BMAL1 transactivation by binding to BMAL1, counteracting its phosphorylation, increasing DNA occupancy, and decreasing transcriptional activity.
Conclusions:
- PPP4 plays a crucial role in regulating circadian rhythm dynamics.
- PPP4 delays negative feedback by modulating CLOCK/BMAL1 activity, thereby fine-tuning the circadian clock.
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