scholarly journals Pacemaker-neuron–dependent disturbance of the molecular clockwork by a Drosophila CLOCK mutant homologous to the mouse Clock mutation

2016 ◽  
Vol 113 (33) ◽  
pp. E4904-E4913 ◽  
Author(s):  
Euna Lee ◽  
Eunjoo Cho ◽  
Doo Hyun Kang ◽  
Eun Hee Jeong ◽  
Zheng Chen ◽  
...  
Keyword(s):  
Clock Genes ◽  
Cellular Level ◽  
Temperature Cycle ◽  
Physical Interaction ◽  
Temperature Transition ◽  
Pacemaker Neurons ◽  
Pacemaker Neuron ◽  
Clock Proteins ◽  
E Box ◽  
Free Running

Circadian clocks are composed of transcriptional/translational feedback loops (TTFLs) at the cellular level. In Drosophila TTFLs, the transcription factor dCLOCK (dCLK)/CYCLE (CYC) activates clock target gene expression, which is repressed by the physical interaction with PERIOD (PER). Here, we show that amino acids (AA) 657–707 of dCLK, a region that is homologous to the mouse Clock exon 19-encoded region, is crucial for PER binding and E-box–dependent transactivation in S2 cells. Consistently, in transgenic flies expressing dCLK with an AA657–707 deletion in the Clock (Clkout) genetic background (p{dClk-Δ};Clkout), oscillation of core clock genes’ mRNAs displayed diminished amplitude compared with control flies, and the highly abundant dCLKΔ657–707 showed significantly decreased binding to PER. Behaviorally, the p{dClk-Δ};Clkout flies exhibited arrhythmic locomotor behavior in the photic entrainment condition but showed anticipatory activities of temperature transition and improved free-running rhythms in the temperature entrainment condition. Surprisingly, p{dClk-Δ};Clkout flies showed pacemaker-neuron–dependent alterations in molecular rhythms; the abundance of dCLK target clock proteins was reduced in ventral lateral neurons (LNvs) but not in dorsal neurons (DNs) in both entrainment conditions. In p{dClk-Δ};Clkout flies, however, strong but delayed molecular oscillations in temperature cycle-sensitive pacemaker neurons, such as DN1s and DN2s, were correlated with delayed anticipatory activities of temperature transition. Taken together, our study reveals that the LNv molecular clockwork is more sensitive than the clockwork of DNs to dysregulation of dCLK by AA657–707 deletion. Therefore, we propose that the dCLK/CYC-controlled TTFL operates differently in subsets of pacemaker neurons, which may contribute to their specific functions.

scholarly journals Shared Components of the FRQ-Less Oscillator and TOR Pathway Maintain Rhythmicity in Neurospora

2021 ◽  
pp. 074873042199994
Author(s):  
Rosa Eskandari ◽  
Lalanthi Ratnayake ◽  
Patricia L. Lakin-Thomas
Keyword(s):  
Circadian Rhythms ◽  
Clock Genes ◽  
Nutrient Sensing ◽  
Mass Spectrometry Analysis ◽  
Growth Responses ◽  
Tor Pathway ◽  
Spectrometry Analysis ◽  
Binding Partner ◽  
Binding Partners ◽  
Free Running

Molecular models for the endogenous oscillators that drive circadian rhythms in eukaryotes center on rhythmic transcription/translation of a small number of “clock genes.” Although substantial evidence supports the concept that negative and positive transcription/translation feedback loops (TTFLs) are responsible for regulating the expression of these clock genes, certain rhythms in the filamentous fungus Neurospora crassa continue even when clock genes ( frq, wc-1, and wc-2) are not rhythmically expressed. Identification of the rhythmic processes operating outside of the TTFL has been a major unresolved area in circadian biology. Our lab previously identified a mutation ( vta) that abolishes FRQ-less rhythmicity of the conidiation rhythm and also affects rhythmicity when FRQ is functional. Further studies identified the vta gene product as a component of the TOR (Target of Rapamycin) nutrient-sensing pathway that is conserved in eukaryotes. We now report the discovery of TOR pathway components including GTR2 (homologous to the yeast protein Gtr2, and RAG C/D in mammals) as binding partners of VTA through co-immunoprecipitation (IP) and mass spectrometry analysis using a VTA-FLAG strain. Reciprocal IP with GTR2-FLAG found VTA as a binding partner. A Δ gtr2 strain was deficient in growth responses to amino acids. Free-running conidiation rhythms in a FRQ-less strain were abolished in Δ gtr2. Entrainment of a FRQ-less strain to cycles of heat pulses demonstrated that Δ gtr2 is defective in entrainment. In all of these assays, Δ gtr2 is similar to Δ vta. In addition, expression of GTR2 protein was found to be rhythmic across two circadian cycles, and functional VTA was required for GTR2 rhythmicity. FRQ protein exhibited the expected rhythm in the presence of GTR2 but the rhythmic level of FRQ dampened in the absence of GTR2. These results establish association of VTA with GTR2, and their role in maintaining functional circadian rhythms through the TOR pathway.

scholarly journals DEC1 Modulates the Circadian Phase of Clock Gene Expression

2008 ◽  
Vol 28 (12) ◽  
pp. 4080-4092 ◽  
Author(s):  
Ayumu Nakashima ◽  
Takeshi Kawamoto ◽  
Kiyomasa K. Honda ◽  
Taichi Ueshima ◽  
Mitsuhide Noshiro ◽  
...  
Keyword(s):  
Gene Expression ◽  
Clock Genes ◽  
Luciferase Reporter ◽  
Constant Darkness ◽  
Mobility Shift ◽  
Behavioral Rhythms ◽  
Circadian Phase ◽  
Clock Gene Expression ◽  
E Box ◽  
E Boxes

ABSTRACT DEC1 suppresses CLOCK/BMAL1-enhanced promoter activity, but its role in the circadian system of mammals remains unclear. Here we examined the effect of Dec1 overexpression or deficiency on circadian gene expression triggered with 50% serum. Overexpression of Dec1 delayed the phase of clock genes such as Dec1, Dec2, Per1, and Dbp that contain E boxes in their regulatory regions, whereas it had little effect on the circadian phase of Per2 and Cry1 carrying CACGTT E′ boxes. In contrast, Dec1 deficiency advanced the phase of the E-box-containing clock genes but not that of the E′-box-containing clock genes. Accordingly, DEC1 showed strong binding and transrepression on the E box, but not on the E′ box, in chromatin immunoprecipitation, electrophoretic mobility shift, and luciferase reporter assays. Dec1 −/− mice showed behavioral rhythms with slightly but significantly longer circadian periods under conditions of constant darkness and faster reentrainment to a 6-h phase-advanced shift of a light-dark cycle. Knockdown of Dec2 with small interfering RNA advanced the phase of Dec1 and Dbp expression, and double knockdown of Dec1 and Dec2 had much stronger effects on the expression of the E-box-containing clock genes. These findings suggest that DEC1, along with DEC2, plays a role in the finer regulation and robustness of the molecular clock.

PERIODIC INHIBITION OF LIVING PACEMAKER NEURONS (I): LOCKED, INTERMITTENT, MESSY, AND HOPPING BEHAVIORS

1991 ◽  
Vol 01 (03) ◽  
pp. 549-581 ◽  
Author(s):  
J. P. SEGUNDO ◽  
E. ALTSHULER ◽  
M. STIBER ◽  
A. GARFINKEL
Keyword(s):  
Nonlinear Dynamics ◽  
Point Process ◽  
Limit Cycles ◽  
Nonlinear Oscillator ◽  
Strange Attractors ◽  
Synaptic Inhibition ◽  
Pacemaker Neurons ◽  
Pacemaker Neuron ◽  
Exhaustive List

This communication is concerned with an embodiment of periodic nonlinear oscillator driving, the synaptic inhibition of one spike-producing pacemaker neuron by another. Data came from a prototypical living synapse. Analyses centered on a prolonged condition between the transients following the onset and cessation of inhibition. Evaluations were guided by point process mathematics and nonlinear dynamics. A rich and exhaustive list of discharge forms, described precisely and canonically, was observed across different inhibitory rates. Previously unrecognized at synapses, most forms were identified with several well known types from nonlinear dynamics. Ordered by decreasing regularities, they were locked, intermittent (including walk-throughs), messy (including erratic and stammerings) and hopping. Each is discussed within physiological and formal contexts. It is conjectured that (i) locked, intermittent and messy forms reflect limit cycles on 2-tori, quasiperiodic orbits and strange attractors, (ii) noise in neurons hovering around threshold contributes to certain intermittent and stammering behaviors, and (iii) hopping either reflects an attractor with several portions or is nonstationary and noise-induced.

Continuous radio telemetry of hypothalamic temperatures from unrestrained animals

1965 ◽  
Vol 20 (2) ◽  
pp. 321-325 ◽  
Author(s):  
Robert O. Rawson ◽  
Jan A. J. Stolwijk ◽  
Hans Graichen ◽  
Robert Abrams
Keyword(s):  
Motor Activity ◽  
Radio Telemetry ◽  
Temperature Cycle ◽  
Body Temperatures ◽  
Night Temperature ◽  
Hypothalamic Temperature ◽  
Temperature Variations ◽  
Free Running ◽  
Hot Environments

A system of radio telemetry has been designed which continuously records body temperatures of unrestrained animals with a resolution of 0.05 C over transmission distances of 100ˑ1,000 ft, permitting observations on free-running animals for indefinite periods of time. Continuous 24-hr recordings were made of hypothalamic temperatures telemetered from cold-acclimatized and unacclimatized dogs living in cold, neutral, and hot environments. During night hours, dogs usually exhibited a decrease in hypothalamic temperature of 0.5ˑ.0 C below daylight levels. Superimposed on the day-night temperature cycle are marked fluctuations of 0.1ˑ0.5 C at a rate of 0.1 C/min. These variations are associated with the level of motor activity, arousal, and with periods of dozing. Shivering in the cold is exhibited even though hypothalamic temperature may be elevated above a level at which no shivering occurs in a neutral environment. spontaneous hypothalamic temperature variations; cold-acclimatized dogs; day-night temperature cycle Submitted on June 3, 1964

Zur Beziehung zwischen Phasenlage und Spontanfrequenz bei der endogenen Tagesperiodik

1963 ◽  
Vol 18 (2) ◽  
pp. 154-157 ◽  
Author(s):  
Klaus Hoffmann
Keyword(s):  
Phase Angle ◽  
Diurnal Rhythm ◽  
Activity Rhythm ◽  
Sustained Oscillation ◽  
Temperature Cycle ◽  
Biological Processes ◽  
Experimental Demonstration ◽  
Free Running ◽  
Spontaneous Frequency ◽  
Free Running Period

The mechanism underlying the endogenous diurnal periodicity of biological processes can be considered a self-sustained oscillation, which can be entrained to an external cycle. In such oscillations the phase-angle of the entrained cycle depends upon the spontaneous frequency (free-running period) of the oscillator.The activity rhythm of lizards kept in constant light, and in a sinusoidal 24-hour temperature cycle, showed entrainment to this cycle. The phase of the entrained rhythm depended on the spontaneous frequency which was expressed in constant conditions occurring immediately before or after the exposure to the extraneous cycle. This is the first experimental demonstration showing the dependence of phase on the free-running period in an endogenous diurnal rhythm.

scholarly journals An NF-κB–driven lncRNA orchestrates colitis and circadian clock

2020 ◽  
Vol 6 (42) ◽  
pp. eabb5202
Author(s):  
Shuai Wang ◽  
Yanke Lin ◽  
Feng Li ◽  
Zifei Qin ◽  
Ziyue Zhou ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Clock Gene ◽  
Clock Genes ◽  
Experimental Colitis ◽  
Circadian Clock Gene ◽  
Its Gene ◽  
E Box ◽  
Direct Binding ◽  
Central Clock ◽  
Clock Protein

We uncover a cycling and NF-κB–driven lncRNA (named Lnc-UC) that epigenetically modifies transcription of circadian clock gene Rev-erbα, thereby linking circadian clock to colitis. Cycling expression of Lnc-UC is generated by the central clock protein Bmal1 via an E-box element. NF-κB activation in experimental colitis transcriptionally drives Lnc-UC through direct binding to two κB sites. Lnc-UC ablation disrupts colonic expressions of clock genes in mice; particularly, Rev-erbα is down-regulated and its diurnal rhythm is blunted. Consistently, Lnc-UC promotes expression of Rev-erbα (a known dual NF-κB/Nlrp3 repressor) to inactivate NF-κB signaling and Nlrp3 inflammasome in macrophages. Furthermore, Lnc-UC ablation sensitizes mice to experimental colitis and abolishes the diurnal rhythmicity in disease severity. Mechanistically, Lnc-UC physically interacts with Cbx1 protein to reduce its gene silencing activity via H3K9me3, thereby enhancing Rev-erbα transcription and expression. In addition, we identify a human Lnc-UC that has potential to promote Rev-erbα expression and restrain inflammations.

scholarly journals E1A-mediated inhibition of myogenesis correlates with a direct physical interaction of E1A12S and basic helix-loop-helix proteins.

1993 ◽  
Vol 13 (8) ◽  
pp. 4714-4727 ◽  
Author(s):  
D A Taylor ◽  
V B Kraus ◽  
J J Schwarz ◽  
E N Olson ◽  
W E Kraus
Keyword(s):  
Gene Transcription ◽  
Specific Gene ◽  
Deletion Mutants ◽  
Physical Interaction ◽  
Direct Physical Interaction ◽  
Amino Terminal ◽  
High Affinity ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Box

The observation that adenovirus E1A gene products can inhibit differentiation of skeletal myocytes suggested that E1A may interfere with the activity of myogenic basic helix-loop-helix (bHLH) transcription factors. We have examined the ability of E1A to mediate repression of the muscle-specific creatine kinase (MCK) gene. Both the E1A12S and E1A13S products repressed MCK transcription in a concentration-dependent fashion. In contrast, amino-terminal deletion mutants (d2-36 and d15-35) of E1A12S were defective for repression. E1A12S also repressed expression of a promoter containing a multimer of the MCK high-affinity E box (the consensus site for myogenic bHLH protein binding) that was dependent, in C3H10T1/2 cells, on coexpression of a myogenin bHLH-VP16 fusion protein. A series of coprecipitation experiments with glutathione S-transferase fusion and in vitro-translated proteins demonstrated that E1A12S, but not amino-terminal E1A deletion mutants, could bind to full-length myogenin and E12 and to deletion mutants of myogenin and E12 that spare the bHLH domains. Thus, the bHLH domains of myogenin and E12, and the high-affinity E box, are targets for E1A-mediated repression of the MCK enhancer, and domains of E1A required for repression of muscle-specific gene transcription also mediate binding to bHLH proteins. We conclude that E1A mediates repression of muscle-specific gene transcription through its amino-terminal domain and propose that this may involve a direct physical interaction between E1A and the bHLH region of myogenic determination proteins.

scholarly journals Clock proteins regulate spatiotemporal organization of clock genes to control circadian rhythms

2020 ◽  
Author(s):  
Yangbo Xiao ◽  
Ye Yuan ◽  
Mariana Jimenez ◽  
Neeraj Soni ◽  
Swathi Yadlapalli
Keyword(s):  
Gene Expression ◽  
Circadian Rhythms ◽  
Nuclear Envelope ◽  
Circadian Clock ◽  
Clock Genes ◽  
Circadian Clocks ◽  
Spatiotemporal Organization ◽  
Clock Proteins ◽  
Expression Behavior ◽  
Clock Protein

ABSTRACTCircadian clocks regulate ∼24 hour oscillations in gene expression, behavior, and physiology. While the molecular and neural mechanisms of circadian rhythms are well characterized, how cellular organization of clock components controls circadian clock regulation remains poorly understood. Here, we elucidate how clock proteins regulate circadian rhythms by controlling the spatiotemporal organization of clock genes. Using high-resolution live imaging techniques we demonstrate that Drosophila clock proteins are concentrated in a few discrete foci and are organized at the nuclear envelope; these results are in contrast to longstanding expectations that clock proteins are diffusely distributed in the nucleus. We also show that clock protein foci are highly dynamic and change in number, size, and localization over the circadian cycle. Further, we demonstrate that clock genes are positioned at the nuclear periphery by the clock proteins precisely during the circadian repression phase, suggesting that subnuclear localization of clock genes plays an important role in the control of rhythmic gene expression. Finally, we show that Lamin B receptor, a nuclear envelope protein, is required for peripheral localization of clock protein foci and clock genes and for normal circadian rhythms. These results reveal that clock proteins form dynamic nuclear foci and play a hitherto unexpected role in the subnuclear reorganization of clock genes to control circadian rhythms, identifying a novel mechanism of circadian regulation. Our results further suggest a new role for clock protein foci in the clustering of clock-regulated genes during the repression phase to control gene co-regulation and circadian rhythms.SIGNIFICANCEAlmost all living organisms have evolved circadian clocks to tell time. Circadian clocks regulate ∼24-hour oscillations in gene expression, behavior and physiology. Here, we reveal the surprisingly sophisticated spatiotemporal organization of clock proteins and clock genes and its critical role in circadian clock function. We show, in contrast to current expectations, that clock proteins are concentrated in a few discrete, dynamic nuclear foci at the nuclear envelope during the repression phase. Further, we uncovered several unexpected features of clock protein foci, including their role in positioning the clock genes at the nuclear envelope precisely during the repression phase to enable circadian rhythms. These studies provide fundamental new insights into the cellular mechanisms of circadian rhythms and establish direct links between nuclear organization and circadian clocks.

scholarly journals Weak Coupling Between Intracellular Feedback Loops Explains Dissociation of Clock Gene Dynamics

2019 ◽  
Author(s):  
Christoph Schmal ◽  
Daisuke Ono ◽  
Jihwan Myung ◽  
J. Patrick Pett ◽  
Sato Honma ◽  
...  
Keyword(s):  
Single Cell ◽  
Clock Genes ◽  
Experimental Studies ◽  
Feedback Loops ◽  
Light Pulses ◽  
Negative Feedback Loops ◽  
Medium Exchange ◽  
A Cell ◽  
Free Running ◽  
Underlying Mechanisms

Circadian rhythms are generated by interlocked transcriptional-translational negative feedback loops (TTFLs), the molecular process implemented within a cell. The contributions, weighting and balancing between the multiple feedback loops remain debated. Dissociated, free-running dynamics in the expression of distinct clock genes has been described in recent experimental studies that applied various perturbations such as slice preparations, light pulses, jet-lag, and culture medium exchange. In this paper, we provide evidence that this "presumably transient" dissociation of circadian gene expression oscillations may occur at the single-cell level. Conceptual and detailed mechanistic mathematical modeling suggests that such dissociation is due to a weak interaction between multiple feedback loops present within a single cell. The dissociable loops provide insights into underlying mechanisms and general design principles of the molecular circadian clock.

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