Regulation of output from the plant circadian clock

Plants, like many other organisms, have endogenous biological clocks that enable them to organize their physiological, metabolic and developmental processes so that they occur at optimal times. The best studied of these biological clocks are the circadian systems that regulate daily (approximately 24 h) rhythms. At the core of the circadian system in every organism are oscillators responsible for generating circadian rhythms. These oscillators can be entrained (set) by cues from the environment, such as daily changes in light and temperature. Completing the circadian clock model are the output pathways that provide a link between the oscillator and the various biological processes whose rhythms it controls. Over the past few years there has been a tremendous increase in our understanding of the mechanisms of the oscillator and entrainment pathways in plants and many useful reviews on the subject. In this review we focus on the output pathways by which the oscillator regulates rhythmic plant processes. In the first part of the review we describe the role of the circadian system in regulation at all stages of a plant's development, from germination and growth to reproductive development as well as in multiple cellular processes. Indeed, the importance of a circadian clock for plants can be gauged by the fact that so many facets of plant development are under its control. In the second part of the review we describe what is known about the mechanisms by which the circadian system regulates these output processes.     

Mmp1 Processing of the PDF Neuropeptide Regulates Circadian Structural Plasticity of Pacemaker Neurons

In the Drosophila brain, the neuropeptide PIGMENT DISPERSING FACTOR (PDF) is expressed in the small and large Lateral ventral neurons (LNvs) and regulates circadian locomotor behavior. Interestingly, PDF immunoreactivity at the dorsal terminals changes across the day as synaptic contacts do as a result of a remarkable remodeling of sLNv projections. Despite the relevance of this phenomenon to circuit plasticity and behavior, the underlying mechanisms remain poorly understood. In this work we provide evidence that PDF along with matrix metalloproteinases (Mmp1 and 2) are key in the control of circadian structural remodeling. Adult-specific downregulation of PDF levels per se hampers circadian axonal remodeling, as it does altering Mmp1 or Mmp2 levels within PDF neurons post-developmentally. However, only Mmp1 affects PDF immunoreactivity at the dorsal terminals and exerts a clear effect on overt behavior. In vitro analysis demonstrated that PDF is hydrolyzed by Mmp1, thereby suggesting that Mmp1 could directly terminate its biological activity. These data demonstrate that Mmp1 modulates PDF processing, which leads to daily structural remodeling and circadian behavior.     

Entrainment of breast (cancer) epithelial cells detects distinct circadian oscillation patterns for clock and hormone receptor genes

Most physiological and biological processes are regulated by endogenous circadian rhythms under the control of both a master clock, which acts systemically and individual cellular clocks, which act at the single cell level. The cellular clock is based on a network of core clock genes, which drive the circadian expression of non-clock genes involved in many cellular processes. Circadian deregulation of gene expression has emerged to be as important as deregulation of estrogen signaling in breast tumorigenesis. Whether there is a mutual deregulation of circadian and hormone signaling is the question that we address in this study. Here we show that, upon entrainment by serum shock, cultured human mammary epithelial cells maintain an inner circadian oscillator, with key clock genes oscillating in a circadian fashion. In the same cells, the expression of the estrogen receptor α (ER A) gene also oscillates in a circadian fashion. In contrast, ER A-positive and -negative breast cancer epithelial cells show disruption of the inner clock. Further, ER A-positive breast cancer cells do not display circadian oscillation of ER A expression. Our findings suggest that estrogen signaling could be affected not only in ER A-negative breast cancer, but also in ER A-positive breast cancer due to lack of circadian availability of ER A. Entrainment of the inner clock of breast epithelial cells, by taking into consideration the biological time component, provides a novel tool to test mechanistically whether defective circadian mechanisms can affect hormone signaling relevant to breast cancer.     

Dexras1: shaping the responsiveness of the circadian clock

Living organisms are endowed with an autonomous timekeeping program that not only maintains circadian rhythms of behaviour and physiology but is reset by cues from the external, cyclic environment. Intracellular signaling events that mediate entrainment of the mammalian circadian clock by photic (light) as well as non-photic inputs are only beginning to be elucidated. Dexras1 is a novel Ras-like G protein that modulates multiple signaling cascades. Genetic ablation of Dexras1 in mice (dexras1(-/-)) results in altered responsiveness of the master circadian clock to photic and non-photic cues. This review will attempt to provide mechanistic insights into the involvement of Dexras1 in biological timing processes based on its role as a modulator of signal transduction.     

Entrainment of breast (cancer) epithelial cells detects distinct circadian oscillation patterns for clock and hormone receptor genes

Most physiological and biological processes are regulated by endogenous circadian rhythms under the control of both a master clock, which acts systemically and individual cellular clocks, which act at the single cell level. The cellular clock is based on a network of core clock genes, which drive the circadian expression of non-clock genes involved in many cellular processes. Circadian deregulation of gene expression has emerged to be as important as deregulation of estrogen signaling in breast tumorigenesis. Whether there is a mutual deregulation of circadian and hormone signaling is the question that we address in this study. Here we show that, upon entrainment by serum shock, cultured human mammary epithelial cells maintain an inner circadian oscillator, with key clock genes oscillating in a circadian fashion. In the same cells, the expression of the estrogen receptor α (ERA) gene also oscillates in a circadian fashion. In contrast, ERA-positive and -negative breast cancer epithelial cells show disruption of the inner clock. Further, ERA-positive breast cancer cells do not display circadian oscillation of ERA expression. Our findings suggest that estrogen signaling could be affected not only in ERA-negative breast cancer, but also in ERA-positive breast cancer due to lack of circadian availability of ERA. Entrainment of the inner clock of breast epithelial cells, by taking into consideration the biological time component, provides a novel tool to test mechanistically whether defective circadian mechanisms can affect hormone signaling relevant to breast cancer.Key words: circadian rhythm, clock genes, estrogen receptor alpha (ERA), breast cancer cells, entrainment, serum shock     

生物发光成像无损伤研究植物生物钟的方法

植物生物钟系统是植物为了适应地球自转进化出的以约24小时为周期的分子系统, 通过感知并整合外界周期性变化的环境信号进而协调细胞内相应基因的表达和能量状态, 赋予植物对生存环境的适应性并参与调控多个植物生长发育过程。目前, 越来越多的研究聚焦于解析植物生物钟的分子机制, 基于此也衍生出很多研究生物钟表型的方法。该文在总结已有生物钟检测方法的基础上, 重点介绍生物钟表型实验中最常用且比较稳定可靠的实验方法, 以期为生物钟的表型研究尤其是生物钟机制研究提供技术支持与借鉴。     

Circadian G2 arrest as related to circadian gating of cell population growth in Euglena

Cell population growth is gated to occur in particular circadian phases, which has been known for over four decades in various organisms including cyanobacteria and human. However, little is known as to which cell cycle phases from G1 to M are primarily regulated by the circadian rhythm or when in a circadian cycle this primary regulation takes place. We report here that in the flagellate alga Euglena gracilis grown photoautotrophically, the circadian rhythm primarily prevented developmentally matured G2 cells from progressing to mitosis, such that cell population growth occurred only during subjective night. In addition, we found that the circadian rhythm also arrests G1-to-S and S-to-G2 transitions at particular circadian phases.     

Gates and oscillators II: zeitgebers and the network model of the brain clock

Circadian rhythms in physiology and behavior are regulated by the SCN. When assessed by expression of clock genes, at least 2 distinct functional cell types are discernible within the SCN: nonrhythmic, light-inducible, retinorecipient cells and rhythmic autonomous oscillator cells that are not directly retinorecipient. To predict the responses of the circadian system, the authors have proposed a model based on these biological properties. In this model, output of rhythmic oscillator cells regulates the activity of the gate cells. The gate cells provide a daily organizing signal that maintains phase coherence among the oscillator cells. In the absence of external stimuli, this arrangement yields a multicomponent system capable of producing a self-sustained consensus rhythm. This follow-up study considers how the system responds when the gate cells are activated by an external stimulus, simulating a response to an entraining (or phase-setting) signal. In this model, the authors find that the system can be entrained to periods within the circadian range, that the free-running system can be phase shifted by timed activation of the gate, and that the phase response curve for activation is similar to that observed when animals are exposed to a light pulse. Finally, exogenous triggering of the gate over a number of days can organize an arrhythmic system, simulating the light-dependent reappearance of rhythmicity in a population of disorganized, independent oscillators. The model demonstrates that a single mechanism (i.e., the output of gate cells) can account for not only free-running and entrained rhythmicity but also other circadian phenomena, including limits of entrainment, a PRC with both delay and advance zones, and the light-dependent reappearance of rhythmicity in an arrhythmic animal.     

Circadian regulation of teleost retinal cone movements in vitro

In the retinas of many species of lower vertebrates, retinal photoreceptors and pigment epithelium pigment granules undergo daily movements in response to both diurnal, and in the case of teleost cone photoreceptors, endogenous circadian signals. Typically, these cone movements take place at dawn and at dusk when teleosts are maintained on a cyclic light (LD) regime, and at expected dawn and expected dusk when animals are maintained in continuous darkness (DD). Because these movements are so strictly controlled, they provide an overt indicator of the stage of the underlying clock mechanism. In this study we report that both light-induced and circadian-driven cone myoid movements in the Midas cichlid (Cichlasoma citrinellum), occur normally in vitro. Many of the features of retinomotor movements found in vivo also occur in our culture conditions, including responses to light and circadian stimuli and dopamine. Circadian induced predawn contraction and maintenance of expected day position in response to circadian modulation, are also normal. Our studies suggest that circadian regulation of cone myoid movement in vitro is mediated locally by dopamine, acting via a D2 receptor. Cone myoid contraction can be induced at midnight and expected mid-day by dark culture with dopamine or the D2 receptor agonist LY171555. Further, circadian induced predawn contraction can be increased with either dopamine or LY171555, or may be reversed with the dopamine D2 antagonist, sulpiride. Sulpiride will also induce cone myoid elongation in retinal cultures at expected mid- day, but will not induce cone myoid elongation at dusk. In contrast, circadian cone myoid movements in vitro were unaffected by the D1 receptor agonist SCH23390, or the D1 receptor antagonist SKF38393. Our short-term culture experiments indicate that circadian regulation of immediate cone myoid movement does not require humoral control but is regulated locally within the retina. The inclusion of dopamine, or dopamine receptor agonists and antagonists in our cultures, has indicated that retinal circadian regulation may be mediated by endogenously produced dopamine, which acts via a D2 mechanism.     

昼夜节律钟调控代谢的研究进展

生理和行为的昼夜节律性调控对健康生活是必需的。越来越多的流行病学和遗传学证据显示昼夜节律的破坏与代谢紊乱性疾病相关联。在分子水平上,昼夜节律受到时钟蛋白组成的转录一翻译负反馈环的调控。时钟蛋白通过以下两种途径调节代谢：首先,时钟蛋白作为转录因子直接调节一些代谢关键步骤的限速酶和代谢相关核受体的表达,其次作为代谢相关核受体的辅调节因子来激活或抑制其转录活性。虽然时钟蛋白对代谢途径的调节导致代谢物水平呈昼夜节律振荡,但是产生的代谢物反过来又可以影响昼夜节律钟基因的表达,进而影响昼夜节律钟。深入研究昼夜节律钟与代谢的交互调节可能为治疗某些代谢紊乱性疾病提供新的治疗方案。     

The pineal gland influences rat circadian activity rhythms in constant light.

Mammalian circadian organization is believed to derive primarily from circadian oscillators within the hypothalamic suprachiasmatic nuclei (SCN). The SCN drives circadian rhythms of a wide array of functions (e.g., locomotion, body temperature, and several endocrine processes, including the circadian secretion of the pineal hormone melatonin). In contrast to the situation in several species of reptiles and birds, there is an extensive literature reporting little or no effect of pinealectomy on mammalian circadian rhythms. However, recent research has indicated that the SCN and circadian systems of several mammalian species are highly sensitive to exogenous melatonin, raising the possibility that endogenous pineal hormone may provide feedback in the control of overt circadian rhythms. To determine the role of the pineal gland in rat circadian rhythms, the effects of pinealectomy on locomotor rhythms in constant light (LL) and constant darkness (DD) were studied. The results indicated that the circadian rhythms of pinealectomized rats but not sham-operated controls dissociated into multiple ultradian components in LL and recoupled into circadian patterns only after 12-21 days in DD. The data suggest that pineal feedback may modulate sensitivity to light and/or provide coupling among multiple circadian oscillators within the SCN.     

Hatching rhythms in the desert locust, Schistocerca gregaria

ABSTRACT. The eggs of Schistocerca gregaria (Orthoptera, Acrididae) incubated under natural conditions hatch only within a few of hours on either side of dawn. This gated hatching is controlled by a circadian clock that is phase set by the diel fluctuations in the temperature of the soil surrounding the eggs. There is a circadian fluctuation in haemolymph sugar concentration which is initiated at least 4 days before hatching. However, eggs hatch arrhythmically unless given cycled temperature incubation for at least 10 days of a 12–13-day incubation. Increase in acetylcholine esterase content of the brain during the penultimate day suggests that increased hatching rhythmicity occurring at this time is the result of increased neural organization. Embryonic activity and respiration show no circadian rhythm but do provide confirmatory evidence of a quiescent phase prior to hatching. This quiescent phase is an integral part of gated hatching behaviour.     

Recording and Analysis of Circadian Rhythms in Running-wheel Activity in Rodents

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day^1-5. Nocturnal rodents, including rats, hamsters, and mice, are active during the night and relatively inactive during the day. Many other behavioral and physiological measures also exhibit daily rhythms, but in rodents, running-wheel activity serves as a particularly reliable and convenient measure of the output of the master circadian clock, the suprachiasmatic nucleus (SCN) of the hypothalamus. In general, through a process called entrainment, the daily pattern of running-wheel activity will naturally align with the environmental light-dark cycle (LD cycle; e.g. 12 hr-light:12 hr-dark). However circadian rhythms are endogenously generated patterns in behavior that exhibit a ~24 hr period, and persist in constant darkness. Thus, in the absence of an LD cycle, the recording and analysis of running-wheel activity can be used to determine the subjective time-of-day. Because these rhythms are directed by the circadian clock the subjective time-of-day is referred to as the circadian time (CT). In contrast, when an LD cycle is present, the time-of-day that is determined by the environmental LD cycle is called the zeitgeber time (ZT).Although circadian rhythms in running-wheel activity are typically linked to the SCN clock^6-8, circadian oscillators in many other regions of the brain and body^9-14 could also be involved in the regulation of daily activity rhythms. For instance, daily rhythms in food-anticipatory activity do not require the SCN^15,16 and instead, are correlated with changes in the activity of extra-SCN oscillators^17-20. Thus, running-wheel activity recordings can provide important behavioral information not only about the output of the master SCN clock, but also on the activity of extra-SCN oscillators. Below we describe the equipment and methods used to record, analyze and display circadian locomotor activity rhythms in laboratory rodents.     

The circadian program of algae

The endogenous circadian program enables organisms to cope with the temporal ecology of their environment. It is driven by a molecular pacemaker, which is found in animals as well as plants at the level of the single cell. Unicellular organisms are, therefore, ideal model systems for the study of circadian systems because rhythms can be investigated in single cells at the molecular, physiological, behavioral and environmental level. In this review, we discuss the possible driving forces for the evolution of circadian rhythmicity in unicellular marine organisms. The current knowledge about the cellular and molecular mechanisms involved in the different components of the circadian system (input, oscillator and output) are described primarily with reference to the marine dinoflagellate,Gonyaulax polyedra. Light is the most important and best described environmental signal synchronizing the endogenous rhythms to the 24-hour solar day. However, little is known about the nature of circadian light receptors, which appear to be distinct from those that control behavioral light responses such as phototaxis. It has recently been shown inGonyaulaxthat nutrients, namely nitrate, can act as a non-photic zeitgeber for the circadian system. In this alga, bioluminescence is under circadian control, and the molecular mechanisms of this circadian output have been investigated in detail. The circadian program turns out to be more complex than simply consisting of an input pathway, a pacemaker and the driven rhythms. Different rhythms appear to be controlled by separate pacemakers, even in single cells, and both circadian inputs and outputs contain feedback loops. The functional advantages of this complexity are discussed. Finally, we outline the differences between the circadian program under laboratory and natural conditions.