scholarly journals Casein Kinase 1 Delta Regulates the Pace of the Mammalian Circadian Clock

2009 ◽  
Vol 29 (14) ◽  
pp. 3853-3866 ◽  
Author(s):  
Jean-Pierre Etchegaray ◽  
Kazuhiko K. Machida ◽  
Elizabeth Noton ◽  
Cara M. Constance ◽  
Robert Dallmann ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Perinatal Period ◽  
Casein Kinase ◽  
Gene Encoding ◽  
Exon 2 ◽  
Casein Kinase 1 ◽  
Functional Differences ◽  
Clock Proteins ◽  
Genes Encoding

ABSTRACT Both casein kinase 1 delta (CK1δ) and epsilon (CK1ε) phosphorylate core clock proteins of the mammalian circadian oscillator. To assess the roles of CK1δ and CK1ε in the circadian clock mechanism, we generated mice in which the genes encoding these proteins (Csnk1d and Csnk1e, respectively) could be disrupted using the Cre-loxP system. Cre-mediated excision of the floxed exon 2 from Csnk1d led to in-frame splicing and production of a deletion mutant protein (CK1δΔ2). This product is nonfunctional. Mice homozygous for the allele lacking exon 2 die in the perinatal period, so we generated mice with liver-specific disruption of CK1δ. In livers from these mice, daytime levels of nuclear PER proteins, and PER-CRY-CLOCK complexes were elevated. In vitro, the half-life of PER2 was increased by ∼20%, and the period of PER2::luciferase bioluminescence rhythms was 2 h longer than in controls. Fibroblast cultures from CK1δ-deficient embryos also had long-period rhythms. In contrast, disruption of the gene encoding CK1ε did not alter these circadian endpoints. These results reveal important functional differences between CK1δ and CK1ε: CK1δ plays an unexpectedly important role in maintaining the 24-h circadian cycle length.

scholarly journals Generation of strong casein kinase 1 inhibitor of Arabidopsis thaliana

2019 ◽  
Author(s):  
Ami N Saito ◽  
Hiromi Matsuo ◽  
Keiko Kuwata ◽  
Azusa Ono ◽  
Toshinori Kinoshita ◽  
...  
Keyword(s):  
Bromine Atom ◽  
Casein Kinase ◽  
In Planta ◽  
Vitro Assay ◽  
Casein Kinase 1 ◽  
Yeast Fungi ◽  
Genes Encoding ◽  
Period Lengthening

AbstractCasein kinase 1 (CK1) is an evolutionarily conserved protein kinase among eukaryotes. Studies on yeast, fungi, and animals have revealed that CK1 plays roles in divergent biological processes. By contrast, the collective knowledge regarding the biological roles of plant CK1 lags was behind those of animal CK1. One of reasons for this is that plants have more multiple genes encoding CK1 than animals. To accelerate the research for plant CK1, a strong CK1 inhibitor that efficiently inhibits multiple members of CK1 proteins in vivo (in planta) is required. Here, we report a novel strong CK1 inhibitor of Arabidopsis (AMI-331). Using a circadian period-lengthening activity as estimation of the CK1 inhibitor effect in vivo, we performed a structure-activity relationship (SAR) study of PHA767491 (1,5,6,7-tetrahydro-2-(4-pyridinyl)-4H-pyrrolo[3,2-c]pyridin-4-one hydrochloride), a potent CK1 inhibitor of Arabidopsis, and found that PHA767491 analogues bearing a propargyl group at the pyrrole nitrogen atom (AMI-212) or a bromine atom at the pyrrole C3 position (AMI-23) enhance the period-lengthening activity. The period lengthening activity of a hybrid molecule of AMI-212 and AMI-23 (AMI-331) is about 100-fold stronger than that of PHA767491. An in vitro assay indicated a strong inhibitory activity of CK1 kinase by AMI-331. Also, affinity proteomics using an AMI-331 probe showed that targets of AMI-331 are mostly CK1 proteins. As such, AMI-331 is a strong potent CK1 inhibitor that shows promise in the research of CK1 in plants.

scholarly journals Casein Kinase 1 Delta (CK1δ) Regulates Period Length of the Mouse Suprachiasmatic Circadian Clock In Vitro

PLoS ONE ◽  
2010 ◽  
Vol 5 (4) ◽  
pp. e10303 ◽  
Author(s):  
Jean-Pierre Etchegaray ◽  
Elizabeth A. Yu ◽  
Premananda Indic ◽  
Robert Dallmann ◽  
David R. Weaver
Keyword(s):  
Circadian Clock ◽  
Casein Kinase ◽  
Period Length ◽  
Casein Kinase 1

Arabidopsis MLK3, a Plant-specific Casein Kinase 1, Negatively Regulated Flowering and Phosphorylated Histone H3 in vivo

2019 ◽  
Author(s):  
Zhen Wang ◽  
Junmei Kang ◽  
Shangang Jia ◽  
Tiejun Zhang ◽  
Zhihai Wu ◽  
...  
Keyword(s):  
Kinase Activity ◽  
Histone H3 ◽  
Casein Kinase ◽  
Kinase Assay ◽  
Wild Type ◽  
Serine Threonine Kinase ◽  
Altered Expression ◽  
Casein Kinase 1 ◽  
Threonine Kinase

Abstract Background: Casein kinase 1 (CK1) family members are highly conserved serine/threonine kinase present in most eukaryotes with multiple biological functions. Arabidopsis MUT9-like kinases ( MLKs ) belong to a clade CK1 specific to the plant kingdom and have been implicated collectively in modulating flowering related processes. Three of the four MLKs ( MLK1/2/4 ) have been characterized, however, little is known about MLK3 , the most divergent MLKs. Results: We demonstrated that compared with wild type, mlk3 , a truncated MLK3 , flowered slightly early under long day conditions and ectopic expression of MLK3 rescued the morphological defects of mlk3 , indicating that MLK3 negatively regulates flowering. GA 3 application accelerated flowering of both wild type and mlk3 , suggesting that mlk3 had normal GA response. The recombinant MLK3-GFP was localized in the nucleus exclusively. In vitro kinase assay revealed that the nuclear protein MLK3 phosphorylated histone 3 at threonine 3 (H3T3ph). Mutation of a conserved catalytic residue (Lysine 175) abolished the kinase activity and resulted in failure to complement the early flowering phenotype of mlk3 . Interestingly, the global level of H3T3 phosphorylation in mlk3 did not differ significantly from wild type, suggesting the redundant roles of MLKs in flowering regulation. The transcriptomic analysis demonstrated that 425 genes significantly altered expression level in mlk3 relative to wild type. The mlk3 mlk4 double mutant generated by crossing mlk3 with mlk4 , a loss-of-function mutant of MLK4 showing late flowering, flowered between the two parental lines, suggesting that MLK3 played an antagonistic role to MLK4 in plant transition to flowering. Conclusions: A serine/threonine kinase encoding gene MLK3 is a casein kinase 1 specific to the plant species and represses flowering slightly. MLK3 located in nucleus catalyzes the phosphorylation of histone H3 at threonine 3 in vitro and an intact lysine residue (K175) is indispensible for the kinase activity. This study sheds new light on the delicate control of flowering by the plant-specific CK1 in Arabidopsis.

scholarly journals CikA, an Input Pathway Component, Senses the Oxidized Quinone Signal to Generate Phase Delays in the Cyanobacterial Circadian Clock

2020 ◽  
Vol 35 (3) ◽  
pp. 227-234 ◽  
Author(s):  
Pyonghwa Kim ◽  
Brianna Porr ◽  
Tetsuya Mori ◽  
Yong-Sung Kim ◽  
Carl H. Johnson ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Time Of Day ◽  
Fine Tuning ◽  
Circadian Oscillation ◽  
Clock Proteins ◽  
Entrainment Process ◽  
Central Oscillator ◽  
External Signals ◽  
Full Synchronization

The circadian clock is a timekeeping system in most organisms that keeps track of the time of day. The rhythm generated by the circadian oscillator must be constantly synchronized with the environmental day/night cycle to make the timekeeping system truly advantageous. In the cyanobacterial circadian clock, quinone is a biological signaling molecule used for entraining and fine-tuning the oscillator, a process in which the external signals are transduced into biological metabolites that adjust the phase of the circadian oscillation. Among the clock proteins, the pseudo-receiver domain of KaiA and CikA can sense external cues by detecting the oxidation state of quinone, a metabolite that reflects the light/dark cycle, although the molecular mechanism is not fully understood. Here, we show the antagonistic phase shifts produced by the quinone sensing of KaiA and CikA. We introduced a new cyanobacterial circadian clock mixture that includes an input component in vitro. KaiA and CikA cause phase advances and delays, respectively, in this circadian clock mixture in response to the quinone signal. In the entrainment process, oxidized quinone modulates the functions of KaiA and CikA, which dominate alternatively at day and night in the cell. This in turn changes the phosphorylation state of KaiC—the central oscillator in cyanobacteria—ensuring full synchronization of the circadian clock. Moreover, we reemphasize the mechanistic input functionality of CikA, contrary to other reports that focus only on its output action.

scholarly journals Synechocystis KaiC3 Displays Temperature- and KaiB-Dependent ATPase Activity and Is Important for Growth in Darkness

2019 ◽  
Vol 202 (4) ◽  
Author(s):  
Anika Wiegard ◽  
Christin Köbler ◽  
Katsuaki Oyama ◽  
Anja K. Dörrich ◽  
Chihiro Azai ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Circadian Clock ◽  
Atp Hydrolysis ◽  
Synechococcus Elongatus ◽  
Constant Darkness ◽  
Content Type ◽  
Clock Proteins ◽  
Pcc 7942 ◽  
Pcc 6803

ABSTRACT Cyanobacteria form a heterogeneous bacterial group with diverse lifestyles, acclimation strategies, and differences in the presence of circadian clock proteins. In Synechococcus elongatus PCC 7942, a unique posttranslational KaiABC oscillator drives circadian rhythms. ATPase activity of KaiC correlates with the period of the clock and mediates temperature compensation. Synechocystis sp. strain PCC 6803 expresses additional Kai proteins, of which KaiB3 and KaiC3 proteins were suggested to fine-tune the standard KaiAB1C1 oscillator. In the present study, we therefore characterized the enzymatic activity of KaiC3 as a representative of nonstandard KaiC homologs in vitro. KaiC3 displayed ATPase activity lower than that of the Synechococcus elongatus PCC 7942 KaiC protein. ATP hydrolysis was temperature dependent. Hence, KaiC3 is missing a defining feature of the model cyanobacterial circadian oscillator. Yeast two-hybrid analysis showed that KaiC3 interacts with KaiB3, KaiC1, and KaiB1. Further, KaiB3 and KaiB1 reduced in vitro ATP hydrolysis by KaiC3. Spot assays showed that chemoheterotrophic growth in constant darkness is completely abolished after deletion of ΔkaiAB1C1 and reduced in the absence of kaiC3. We therefore suggest a role for adaptation to darkness for KaiC3 as well as a cross talk between the KaiC1- and KaiC3-based systems. IMPORTANCE The circadian clock influences the cyanobacterial metabolism, and deeper understanding of its regulation will be important for metabolic optimizations in the context of industrial applications. Due to the heterogeneity of cyanobacteria, characterization of clock systems in organisms apart from the circadian model Synechococcus elongatus PCC 7942 is required. Synechocystis sp. strain PCC 6803 represents a major cyanobacterial model organism and harbors phylogenetically diverged homologs of the clock proteins, which are present in various other noncyanobacterial prokaryotes. By our in vitro studies we unravel the interplay of the multiple Synechocystis Kai proteins and characterize enzymatic activities of the nonstandard clock homolog KaiC3. We show that the deletion of kaiC3 affects growth in constant darkness, suggesting its involvement in the regulation of nonphotosynthetic metabolic pathways.

scholarly journals Real-Time In Vitro Fluorescence Anisotropy of the Cyanobacterial Circadian Clock

2019 ◽  
Vol 2 (2) ◽  
pp. 42 ◽  
Author(s):  
Joel Heisler ◽  
Archana Chavan ◽  
Yong-Gang Chang ◽  
Andy LiWang
Keyword(s):  
Circadian Clock ◽  
Real Time ◽  
Fluorescence Anisotropy ◽  
Polyacrylamide Gel Electrophoresis ◽  
Live Cells ◽  
Ex Post ◽  
Clock Proteins ◽  
Ex Post Facto ◽  
Clock Reaction

Uniquely, the circadian clock of cyanobacteria can be reconstructed outside the complex milieu of live cells, greatly simplifying the investigation of a functioning biological chronometer. The core oscillator component is composed of only three proteins, KaiA, KaiB, and KaiC, and together with ATP they undergo waves of assembly and disassembly that drive phosphorylation rhythms in KaiC. Typically, the time points of these reactions are analyzed ex post facto by denaturing polyacrylamide gel electrophoresis, because this technique resolves the different states of phosphorylation of KaiC. Here, we describe a more sensitive method that allows real-time monitoring of the clock reaction. By labeling one of the clock proteins with a fluorophore, in this case KaiB, the in vitro clock reaction can be monitored by fluorescence anisotropy on the minutes time scale for weeks.

scholarly journals Autokinase Activity of Casein Kinase 1 δ/ε Governs the Period of Mammalian Circadian Rhythms

2019 ◽  
Vol 34 (5) ◽  
pp. 482-496 ◽  
Author(s):  
Gaili Guo ◽  
Kankan Wang ◽  
Shan-Shan Hu ◽  
Tian Tian ◽  
Peng Liu ◽  
...  
Keyword(s):  
Circadian Rhythms ◽  
Conformational Changes ◽  
Catalytic Domain ◽  
Biochemical Properties ◽  
Period Change ◽  
Casein Kinase ◽  
Full Length ◽  
Casein Kinase 1 ◽  
Autokinase Activity

Circadian rhythms exist in nearly all organisms. In mammals, transcriptional and translational feedback loops (TTFLs) are believed to underlie the mechanism of the circadian clock. Casein kinase 1δ/ε (CK1δ/ε) are key kinases that phosphorylate clock components such as PER proteins, determining the pace of the clock. Most previous studies of the biochemical properties of the key kinases CK1ε and CK1δ in vitro have focused on the properties of the catalytic domains from which the autoinhibitory C-terminus has been deleted (ΔC); those studies ignored the significance of self-inhibition by autophosphorylation. By comparing the properties of the catalytic domain of CK1δ/ε with the full-length kinase that can undergo autoinhibition, we found that recombinant full-length CK1 showed a sequential autophosphorylation process that induces conformational changes to affect the overall kinase activity. Furthermore, a direct relationship between the period change and the autokinase activity among CK1δ, CK1ε, and CK1ε-R178C was observed. These data implicate the autophosphorylation activity of CK1δ and CK1ε kinases in setting the pace of mammalian circadian rhythms and indicate that the circadian period can be modulated by tuning the autophosphorylation rates of CK1δ/ε.

scholarly journals Arabidopsis thaliana MLK3, a Plant-Specific Casein Kinase 1, Negatively Regulates Flowering and Phosphorylates Histone H3 In Vitro

Genes ◽  
2020 ◽  
Vol 11 (3) ◽  
pp. 345 ◽  
Author(s):  
Junmei Kang ◽  
Huiting Cui ◽  
Shangang Jia ◽  
Wenwen Liu ◽  
Renjie Yu ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Flowering Time ◽  
Leaf Growth ◽  
Ectopic Expression ◽  
Casein Kinase ◽  
Kinase Assay ◽  
Casein Kinase 1 ◽  
Histone 3 ◽  
Morphological Defects

Arabidopsis thaliana MUT9-LIKE KINASES (MLKs), a family of the plant-specific casein kinase 1 (CK1), have been implicated collectively in multiple biological processes including flowering. Three of the four MLKs (MLK1/2/4) have been characterized, however, little is known about MLK3, the most divergent member of MLKs. Here, we demonstrated that disruption of MLK3 transcript in mlk3 caused early flowering with retarded leaf growth under long-day conditions. In vitro kinase assay showed the nuclear protein MLK3 phosphorylated histone 3 at threonine 3 (H3T3) and mutation of a conserved residue (K146R) abolished the catalytic activity. Ectopic expression of MLK3 but not MLK3(K146R) rescued the morphological defects of mlk3, indicating that an intact MLK3 is critical for maintaining proper flowering time. Transcriptomic analysis revealed that the floral repressor FLOWERING LOCUS C (FLC) was down-regulated significantly in mlk3, suggesting that MLK3 negatively regulates flowering. Hence, MLK3 plays a role in repressing the transition from vegetative to reproductive phase in A. thaliana. This study sheds light on the delicate control of flowering time by A. thaliana CK1 specific to the plant kingdom.

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