Dataset Information

CK1?/? protein kinase primes the PER2 circadian phosphoswitch.

ABSTRACT: Multisite phosphorylation of the PERIOD 2 (PER2) protein is the key step that determines the period of the mammalian circadian clock. Previous studies concluded that an unidentified kinase is required to prime PER2 for subsequent phosphorylation by casein kinase 1 (CK1), an essential clock component that is conserved from algae to humans. These subsequent phosphorylations stabilize PER2, delay its degradation, and lengthen the period of the circadian clock. Here, we perform a comprehensive biochemical and biophysical analysis of mouse PER2 (mPER2) priming phosphorylation and demonstrate, surprisingly, that CK1?/? is indeed the priming kinase. We find that both CK1? and a recently characterized CK1?2 splice variant more efficiently prime mPER2 for downstream phosphorylation in cells than the well-studied splice variant CK1?1. While CK1 phosphorylation of PER2 was previously shown to be robust to changes in the cellular environment, our phosphoswitch mathematical model of circadian rhythms shows that the CK1 carboxyl-terminal tail can allow the period of the clock to be sensitive to cellular signaling. These studies implicate the extreme carboxyl terminus of CK1 as a key regulator of circadian timing.

SUBMITTER: Narasimamurthy R

PROVIDER: S-EPMC6003379 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

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CK1δ/ε protein kinase primes the PER2 circadian phosphoswitch.

Narasimamurthy Rajesh R Hunt Sabrina R SR Lu Yining Y Fustin Jean-Michel JM Okamura Hitoshi H Partch Carrie L CL Forger Daniel B DB Kim Jae Kyoung JK Virshup David M DM

Proceedings of the National Academy of Sciences of the United States of America 20180521 23

Multisite phosphorylation of the PERIOD 2 (PER2) protein is the key step that determines the period of the mammalian circadian clock. Previous studies concluded that an unidentified kinase is required to prime PER2 for subsequent phosphorylation by casein kinase 1 (CK1), an essential clock component that is conserved from algae to humans. These subsequent phosphorylations stabilize PER2, delay its degradation, and lengthen the period of the circadian clock. Here, we perform a comprehensive bioch ...[more]

PMID: 29784789

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Project description:Like neurons in the suprachiasmatic nucleus (SCN), the master circadian pacemaker in the brain, single fibroblasts can function as independent oscillators. In the SCN, synaptic and paracrine signaling among cells creates a robust, synchronized circadian oscillation, whereas there is no evidence for such integration in fibroblast cultures. However, interactions among single-cell fibroblast oscillators cannot be completely excluded, because fibroblasts were not isolated in previous work. In this study, we tested the autonomy of fibroblasts as single-cell circadian oscillators in high- and low-density culture, by single-cell imaging of cells from PER2::LUC circadian reporter mice. We found greatly reduced PER2::LUC rhythmicity in low-density cultures, which could result from lack of either constitutive or rhythmic paracrine signals from neighboring fibroblasts. To discriminate between these 2 possibilities, we mixed PER2::LUC wild-type (WT) cells with nonluminescent, nonrhythmic Bmal1-/- cells, so that density of rhythmic cells was low but overall cell density remained high. In this condition, WT cells showed clear rhythmicity similar to high-density cultures. We also mixed PER2::LUC WT cells with nonluminescent, long period Cry2-/- cells. In this condition, WT cells showed a period no different from cells cultured with rhythmic WT cells or nonrhythmic Bmal1-/- cells. In previous work, we found that low K? suppresses fibroblast rhythmicity, and we and others have found that either low K? or low Ca²? suppresses SCN rhythmicity. Therefore, we attempted to rescue rhythmicity of low-density fibroblasts with high K? (21 mM), high Ca²? (3.6 mM), or conditioned medium. Conditioned medium from high-density fibroblast cultures rescued rhythmicity of low-density cultures, whereas high K? or Ca²? medium did not consistently rescue rhythmicity. These data suggest that fibroblasts require paracrine signals from adjacent cells for normal expression of rhythmicity, but that these signals do not have to be rhythmic, and that rhythmic signals from other cells do not affect the intrinsic periods of fibroblasts.

Dataset Information

CK1?/? protein kinase primes the PER2 circadian phosphoswitch.

Publications

CK1δ/ε protein kinase primes the PER2 circadian phosphoswitch.

Similar Datasets

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