The circadian conidiation rhythm inNeurospora crassa

The filamentous fungusNeurospora crassais one of the best organisms for analysing the molecular basis of the circadian rhythm observed in asexual spore formation, conidiation. Many clock mutants in which the circadian conidiation rhythm has different characteristics compared to those in the wild-type strain have been isolated since the early 1970s. With the cloning of one of these clock genes,frq, the molecular basis of the circadian clock inNeurosporahas become gradually clearer. Physiological and pharmacological studies have also contributed to our understanding of the physiological basis of the circadian clock inNeurospora. These studies strongly indicate that the circadian clock is based on or is closely related to a network of metabolic processes for cellular activities. Based on these studies, it may be possible to isolate new types of clock mutants which should contribute to a better understanding of the molecular basis of the circadian clock inNeurospora.     

Egg-laying rhythm in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

Extensive research has been carried out to understand how circadian clocks regulate various physiological processes in organisms. The discovery of clock genes and the molecular clockwork has helped researchers to understand the possible role of these genes in regulating various metabolic processes. In Drosophila melanogaster, many studies have shown that the basic architecture of circadian clocks is multi-oscillatory. In nature, different neuronal subgroups in the brain of D. melanogaster have been demonstrated to control different circadian behavioural rhythms or different aspects of the same circadian rhythm. Among the circadian phenomena that have been studied so far in Drosophila, the egg-laying rhythm is unique, and relatively less explored. Unlike most other circadian rhythms, the egg-laying rhythm is rhythmic under constant light conditions, and the endogenous or free-running period of the rhythm is greater than those of most other rhythms. Although the clock genes and neurons required for the persistence of adult emergence and activity/rest rhythms have been studied extensively, those underlying the circadian egg-laying rhythm still remain largely unknown. In this review, we discuss our current understanding of the circadian egg-laying rhythm in D. melanogaster, and the possible molecular and physiological mechanisms that control the rhythmic output of the egg-laying process.     

自养和兼养条件下蛋白核小球藻昼夜节律的响应

【目的】研究自养和兼养两种培养方式对蛋白核小球藻(Chlorella pyrenoidosa)生长、细胞分裂和生化组分积累的影响,探讨人工培养蛋白核小球藻的昼夜节律响应机制和优化技术。【方法】小球藻自养培养采用BG11培养基,兼养培养基在BG11培养基中添加4种不同浓度(1、5、10、20 g/L)的葡萄糖,培养周期为10 d。血球板计数法测定藻细胞浓度,干重法测定藻细胞生物量。显微观察藻细胞大小和分裂情况。脂染色法测定小球藻总脂的含量,藻细胞的叶绿素、蛋白和淀粉分别采用甲醇、氢氧化钠、硝酸钙浸提后通过紫外分光光度法定量测定。【结果】葡萄糖兼养培养对蛋白核小球藻具有显著的促生长效应,最适浓度为10 g/L。10 d收获时,兼养组(10 g/L葡萄糖)藻细胞浓度和干重分别是自养组的2.57倍和6.73倍。分析一昼夜中的藻细胞增殖规律可知,第2天和第5天时自养组中增殖的新生子细胞约有76.00%在黑暗期分裂产生,而兼养组中第2天和第5天光照期的新细胞增殖量占比分别达到40.90%和67.50%。一昼夜内藻细胞大小的迁移动态监测表明,第2天自养组藻细胞的体积变化静息期为8 h,兼养组只有4 h;第5天两组藻细胞大小迁移动态的昼夜节律明显,但兼养组黑暗结束后较大细胞(D6μm)占比显著高于自养组。第8天时,兼养组藻细胞已处于稳定期,总脂和蛋白含量均显著高于自养组,藻细胞总脂和色素含量在一昼夜中相对稳定,但蛋白和淀粉含量分别在光照8 h和12 h左右达到峰值。从第2天开始,对兼养组细胞每天进行2 h光延长,收获时藻细胞浓度和干重分别比对照组提高13%和11%。【结论】葡萄糖兼养培养能大幅提高蛋白核小球藻的生物量。蛋白核小球藻生长增殖与生化组分积累均受昼夜节律调控,自养条件下藻细胞以光照期生长黑暗期增殖为主。兼养培养提高藻细胞生物量的机制在于缩短藻细胞生长静息期,在昼夜节律中加速藻细胞生长并显著提高通过细胞周期检查点的细胞比例,光照期效应尤其明显。藻细胞蛋白和淀粉含量昼夜节律明显,最佳收获时间分别在光照8 h和12 h后。     

A circadian clock measures photoperiodic time in the male lizard Anolis carolinensis

Summary Photoperiod plays an important role in controlling the annual reproductive cycle of the male lizard Anolis carolinensis. The nature of photoperiodic time measurement in Anolis was investigated by exposing anoles to 3 different kinds of lighting paradigms (resonance, T cycles, and night breaks) to determine if photoperiodic time measurement involves the circadian system. Both the reproductive response and the patterns of entrainment of the activity rhythm were assessed. The results show that the circadian system is involved in photoperiodic time measurement in this species and that a discrete photoinducible phase resides in the latter half of the animals' subjective night. Significantly, the ability of the circadian system to execute photoperiodic time measurement is crucially dependent on the length of the photoperiod. Resonance, T cycle and night break cycles utilizing a photoperiod 10–11 h in duration reveal circadian involvement whereas these same cycles utilizing 6 or 8 h photoperiods do not.Abbreviation CRPP circadian rhythm of photoperiodic sensitivity     

Automated Recording of Vertical Negative Phototactic Behaviour in Daphnia magna Straus (Crustacea)

Diurnal vertical migration is a well-known phenomenon in the circadian activity rhythms of zooplankton. Our goal was to test whether negative phototaxis in Daphnia magna clone BEAK (provoked by artificially induced light stress, alternating light and dark phases in 2 h intervals), and its interference with the endogenous rhythm of diurnal vertical migration, can be automatically registered with a biomonitor. For the first time the vertical swimming behaviour of D. magna was recorded quantitatively based on non-optical data recording in a fully automated biotest system, the Multispecies Freshwater Biomonitor in a new experimental setup consisting of a column of three recording units (3-level chambers). Circadian vertical migration was clearly recorded in the 3-level chambers and the rhythm was more clear with 5 than with 1 organism per chamber. The organisms clearly responded to induced light stress with negative phototaxis, however best in larger chambers. The artificially induced rhythm was influenced by the endogenous rhythm. This approach may facilitate long-term observations of vertical swimming activity of zooplankton in the future.     

Light responsiveness of clock genes, Per1 and Per2, in the olfactory bulb of mice

The olfactory bulb (OB) of rodents has been suggested to possess a self-sustaining circadian oscillator which functions independent from the master circadian clock in the suprachiasmatic nucleus (SCN) of the hypothalamus. However, neither histology nor physiology of this extra-SCN clock is studied yet. In the present study, we examined circadian variation of major clock gene expressions in the OB and responsiveness to single photic stimuli. Here we show significant circadian variation in the expression of clock genes, Per1, Per2 and Bmal1 in the OB. Per1 and PER2 were mainly expressed in the mitral cell and granular cell layers of the OB. Light responsiveness of Per1 and Per2 expression was different in the OB from that in the parietal cortex. Both Per1 and Per2 are expressed in the OB only by l000 lux light pulse, whereas 100 lux light was enough to induce Per1 mRNA in the parietal cortex. Interestingly, even 1000 lux light failed to induce Per2 mRNA in the parietal cortex. These clock gene-specific and brain region-dependent responses to lights in the OB and parietal cortex suggest that single light stimulus induces various physiological functions in different brain areas via specific clock gene.     

模式生物粗糙脉孢菌光受体的研究进展

粗糙脉孢菌是一种重要的模式生物,在遗传调节机制、昼夜节律运行以及真菌光应答反应研究中起重要的作用.本综述主要介绍粗糙脉孢菌光受体WC-1和VVD的结构与功能,以及它们参与调节昼夜节律和光适应机制方面的研究进展.在该真菌中,所有已知的光应答反应都受蓝光调节,由光受体WC-1和VVD介导.WC-1是该真菌的转录因子,介导最初的光反应过程,产生VVD等多种光反应蛋白,而VVD通过负反馈机制抑制WC-1的转录作用.此外,vvd基因已经用于构建在哺乳动物中表达的光调节基因元件.     

The role of temperature in the regulation of the circadian rhythm of CO2 fixation in Bryophyllum fedtschenkoi

Detached leaves of Bryophyllum fedtschenkoi Hamet et Perrier kept in normal air show a single period of net CO₂ fixation on transfer to constant darkness at temperatures in the range 0–25 °C. The duration of this initial fixation period is largely independent of temperature in the range 5–20 °C, but lengthens very markedly at temperatures below 4 °C, and is reduced at temperatures above 25 °C. The onset of net fixation of CO₂ on transfer of leaves to constant darkness is immediate at low temperatures, but is delayed as the temperature is increased. The ambient temperature also determines whether or not a circadian rhythm of CO₂ exchange occurs. The rhythm begins to appear at about 20 °C, is most evident at 30 °C and becomes less distinct at 35 °C. The occurrence of a distinct circadian rhythm in CO₂ output at 30° C in the absence of a detectable rhythm in PEPCase kinase activity shows that the kinase rhythm is not a mandatory requirement for the rhythm of PEPCase activity. However, when it occurs, the kinase rhythm undoubtedly amplifies the PEPCase rhythm.Abbreviation PEPCase phosphoenolpyruvate carboxylase We thank the Agricultural and Food Research Council for financial support for this work.     

Circadian rhythms of locomotor activity and temperature selection in sleepy lizards, <Emphasis Type="Italic">Tiliqua rugosa</Emphasis>

This study examined whether the daily rhythms of locomotor activity and behavioural thermoregulation that have previously been observed in Australian sleepy lizards (Tiliqua rugosa) under field conditions are true circadian rhythms that persist in constant darkness (DD) and whether these rhythms show similar characteristics. Lizards held on laboratory thermal gradients in the Australian spring under the prevailing 12-hour light : dark (LD) cycle for 14 days displayed robust daily rhythms of behavioural thermoregulation and locomotor activity. In the 13-day period of DD that followed LD, most lizards exhibited free-running circadian rhythms of locomotor activity and behavioural thermoregulation. The predominant activity pattern displayed in LD was unimodal and this was retained in DD. While mean levels of skin temperature and locomotor activity were found to decrease from LD to DD, activity duration remained unchanged. The present results demonstrate for the first time that this species’ daily rhythm of locomotor activity is an endogenous circadian rhythm. Our results also demonstrate a close correlation between the circadian activity and thermoregulatory rhythms in this species indicating that the two rhythms are controlled by the same master oscillator(s). Future examination of seasonal aspects of these rhythms, may, however, cause this hypothesis to be modified.     

Correlation between circadian periods and cellular activities in Paramecium bursaria

Paramecium bursaria shows a circadian rhythm of photoaccumulation: photoaccumulation is stronger during the day than at night. We obtained five strains of P. bursaria having different circadian periods under continuous light conditions, ranging from 20.9 to 27.9 h. Various physiological activities were compared in the cells of these strains. The periods of contractile vacuole contraction were in the range 10–15 s, which was almost proportional to the periods of the circadian rhythm in each strain. Swimming velocities were inversely proportional to the circadian period; i.e. swimming velocities were high in strains whose circadian periods were short. Resting membrane potential was more depolarized in strains with longer circadian periods. Finally, the membrane resistance of the resting state was reduced in proportion to the increase of the circadian period. Such correlation between the cellular properties and the circadian period suggests that the circadian clock mechanism is associated with various physiological activities of the cell.     

hormone nuclear receptor (EcR) exhibits circadian cycling in certain tissues, but not others, during development in Rhodnius prolixus (Hemiptera)

The insect moulting hormones, viz. the ecdysteroids, regulate gene expression during development by binding to an intracellular protein, the ecdysteroid receptor (EcR). In the insect Rhodnius prolixus, circulating levels of ecdysteroids exhibit a robust circadian rhythm. This paper demonstrates associated circadian rhythms in the abundance and distribution of EcR in several major target tissues of ecdysteroids, but not in others. Quantitative analysis of immunofluorescence images obtained by confocal laser-scanning microscopy following the use of anti-EcR has revealed a marked daily rhythm in the nuclear abundance of EcR in cells of the abdominal epidermis, brain, fat body, oenocytes and rectal epithelium of Rhodnius. This EcR rhythm is synchronous with the rhythm of circulating hormone levels. It free-runs in continuous darkness for several cycles, showing that EcR nuclear abundance is under circadian control. Circadian control of a nuclear receptor has not been shown previously in any animal. We infer that the above cell types detect and respond to the temporal signals in the rhythmic ecdysteroid titre. In several cell types, the rhythm in cytoplasmic EcR peaks several hours prior to the EcR peak in the nucleus each day, thereby implying a daily migration of EcR from the cytoplasm to the nucleus. This finding shows that EcR is not a constitutive nuclear receptor, as has previously been assumed. In the brain, rhythmic nuclear EcR has been found in peptidergic neurosecretory cells, indicating a potential pathway for feedback regulation of the neuroendocrine system by ecdysteroids, and also in regions containing circadian clock neurons, suggesting that the circadian timing system in the brain is also sensitive to rhythmic ecdysteroid signals. This work was supported by a Discovery Grant from the Natural Sciences and Engineering Research Council of Canada.     

Circadian rhythm of (Z)- and (E)-2-β-d-glucopyranosyloxy-4-methoxy cinnamic acids and herniarin in leaves of Matricaria chamomilla

Chamomile (Matricaria chamomilla) in the above-ground organs synthesizes and accumulates (Z)- and (E)-2-β-d-glucopyranosyloxy-4-methoxy cinnamic acids (GMCA), the precursors of phytoanticipin herniarin (7-methoxycoumarin). The diurnal rhythmicity of the sum of GMCA (maximum before daybreak) and herniarin (acrophase at 10 h 21 min of circadian time) was observed under artificial lighting conditions LD 12:12. The acrophase is the time point of the maximum of the sinusoidal curve fitted to the experimental data. In continuous light, the circadian rhythms of both compounds were first described with similar acrophases of endogenous rhythms; a significantly different result from that in synchronized conditions. The rhythms’ mesor (the mean value of the sinusoidal curve fitted to the experimental data) under free-running conditions was not influenced. Abiotic stress under synchronized conditions decreased the average content of GMCA to half of the original level and eliminated the rhythmicity. In contrast, the rhythm of herniarin continued, though its content significantly increased. Nitrogen deficiency resulted in a significant increase in GMCA content, which did not manifest any rhythmicity while the rhythm of herniarin continued. Circadian control of herniarin could be considered as a component of the plant’s specialized defence mechanisms.     

Circadian control of heat tolerance in stationary phase cultures of Schizosaccharomyces pombe

The capacity of stationary phase cultures of Schizosaccharomyces pombe to survive a heat treatment at 55°C is controlled by a circadian rhythm. In a synchronizing light-dark-cycle this rhythm shows a stable phase relationship to the onset of light. In continuous darkness it persists for several cycles without marked damping. The free-running period of about 27 h at 30°C is only slightly longer at 20°C, hence temperature-compensated. These results indicate that S. pombe is a suitable experimental organism for further research into both heat tolerance and circadian rhythms.     

Locomotor activity rhythm and sun compass orientation in the sandhopper <Emphasis Type="Italic">Talitrus </Emphasis><Emphasis Type="Italic">saltator</Emphasis> are related

The sandhopper Talitrus saltator has an endogenous activity rhythm with a circadian periodicity. It is well known for its ability to compensate for the apparent movement of the sun during its migrations along the sea–land axis of the beach. Both chronometric mechanisms are entrained by the natural LD photoperiod. Using actographic recordings and tests of solar orientation of individuals kept under an LD 12:12 clock-shifted cycle, after 1–13 days of treatment, we demonstrate that the timing mechanism of activity rhythm and the chronometric mechanism underlying the sun compass are the same.     

Perturbations of malate accumulation and the endogenous rhythms of gas exchange in the Crassulacean acid metabolism plant<Emphasis Type="Italic"> Kalanchoë daigremontiana</Emphasis>: testing the tonoplast-as-oscillator model

In continuous light, leaves of the Crassulacean acid metabolism (CAM) plant Kalanchoë daigremontiana Hamet et Perrier exhibit a circadian rhythm of CO₂ uptake, stomatal conductance and leaf-internal CO₂ pressure. According to a current quantitative model of CAM, the pacemaking mechanism involves periodic turgor-related tension and relaxation of the tonoplast, which determines the direction of the net flux of malate between the vacuole and the cytoplasm. Cytoplasmic malate, in turn, through its inhibitory effect on phosphoenolpyruvate carboxylase, controls the rate of CO₂ uptake. According to this mechanism, when the accumulation of malate is disrupted by removing CO₂ from the ambient air, the induction of a phase delay with respect to an unperturbed control plant is expected. First, using the mathematical model, such phase delays were observed in numerical simulations of three scenarios of CO₂ removal: (i) starting at a trough of CO₂ uptake, lasting for about half a cycle (ca. 12 h in vivo); (ii) with the identical starting phase, but lasting for 1.5 cycles (ca. 36 h); and (iii) starting while CO₂ increases, lasting for half a cycle again. Applying the same protocols to leaves of K. daigremontiana in vivo did not induce the predicted phase shifts, i.e. after the end of the CO₂ removal the perturbed rhythm adopted nearly the same phase as that of the control plant. Second, when leaves were exposed to a nitrogen atmosphere for three nights prior to onset of continuous light to prevent malate accumulation, a small, 4-h phase advance was observed instead of a delay, again contrary to the model-based expectations. Hence, vacuolar malic acid accumulation is ruled out as the central pacemaking process. This observation is in line with our earlier suggestion [T.P. Wyka, U. Lüttge (2003) J Exp Bot 54:1471–1479] that in extended continuous light, CO₂ uptake switches gradually from a CAM-like to a C3-like mechanism, with oscillations of the two CO₂ uptake systems being tightly coordinated. It appears that the circadian rhythm of gas exchange in this CAM plant emerges from one or several devices that are capable of generating temporal information in a robust manner, i.e. they are protected from even severe metabolic perturbations.Abbreviations CAM Crassulacean acid metabolism - c_ia Ratio of mesophyll CO₂ concentration to external CO₂ concentration - J_C Rate of carbon dioxide uptake - J_W Transpiration rate - g_W Stomatal conductance - LL Continuous light conditions - PEPC Phosphoenolpyruvate carboxylase - Rubisco d-Ribulose-1,5-bisphosphatecarboxylase/oxygenase - Effective quantum yield of photosystem II     

Control of the circadian rhythm of carbon dioxide assimilation in Bryophyllum leaves by exposure to darkness and high carbon dioxide concentrations

The circadian rhythm of CO₂ assimilation in detached leaves of Bryophyllum fedtschenkoi at 15° C in normal air and continuous illumination is inhibited both by exposure to darkness, and to an atmosphere enriched with 5% CO₂. During such exposures substantial fixation of CO₂ takes place, and the malate concentration in the cell sap increases from about 20 mM to a constant value of 40–50 mM after 16 h. On transferring the darkened leaves to light, and those exposed to 5% CO₂ to normal air, a circadian rhythm of CO₂ assimilation begins again. The phase of this rhythm is determined by the time the transfer is made since the first peak occurs about 24 h afterwards. This finding indicates that the circadian oscillator is driven to, and held at, an identical, fixed phase point in its cycle after 16 h exposure to darkness or to 5% CO₂, and it is from this phase point that oscillation begins after the inhibiting condition is removed. This fixed phase point is characterised by the leaves having acquired a high malate content. The rhythm therefore begins with a period of malate decarboxylation which lasts for about 8 h, during which time the malate content of the leaf cells must be reduced to a value that allows phosphoenolpyruvate carboxylase to become active. Inhibition of the rhythm in darkness, and on exposure to 5% CO₂ in continuous illumination, appears to be due to the presence of a high concentration of CO₂ within the leaf inhibiting malic enzyme which leads to the accumulation of high concentrations of malate in the leaf cells. The malate then allosterically inhibits phosphoenolpyruvate carboxylase upon which the rhythm depends. The results give support to the view that malate synthesis and breakdown form an integral part of the circadian oscillator in this tissue.Abbreviations B. Bryophyllum - PEPCase phosphoenolpyruvate carboxylase