生物钟基因研究新进展

生物钟基因普遍存在于生物界,其作用在于产生和控制昼夜节律的运转。生物钟基因及其编码的蛋白质组成反馈回路,维持振荡系统持续进行并与环境周期保持同步。各级进化水平物种生物钟的基因组成和控制途径有同有异。此文主要介绍蓝细菌、脉孢菌、果蝇、鼠和人昼夜钟的分子运作机制以及研究钟基因的意义和展望。 Abstract:The circadian clock genes,which generate and control the running of the circadian rhythms,exist in organisms ranging from prokaryotes to mammals.The oscillator genes and its coding proteins compose the feedback loops of circadian system.The kind,number and regulating route of clock genes are characterized by living things at different evolution levels.The molecular mechanism of the run of circadian clock genes in cyanobacteria,neurospore,fruit fly,mouse and human being is introduced in this article.     

Circadian rhythms of cyanobacteria: monitoring the biological clocks of individual colonies by bioluminescence.

Reproducible circadian rhythms of bioluminescence from individual colonies of cyanobacteria (Synechococcus sp. strain PCC 7942) has been observed. Phenotypic monitoring of colonies on agar plates will enable us to genetically analyze the molecular mechanism of the circadian clock of cyanobacteria by screening for clock mutants. By the introduction of a bacterial luciferase gene, we previously developed a transformed cyanobacterial strain (AMC149) that expresses luciferase as a bioluminescent reporter of the circadian clock. In liquid culture, AMC149 expresses a rhythm of bioluminescence that displays the same behavior as circadian rhythms in higher eukaryotes. Improvements in the technique for administering the reporter enzyme's substrate (decanal) and a highly sensitive photon-counting camera allow monitoring the bioluminescence of single colonies. Individual colonies on agar plates displayed a rhythmicity which is essentially the same as that previously reported for liquid cultures.     

The circadian clock of cyanobacteria

A circadian clock, with physiological characteristics similar to those of eukaryotes, functions in the photosynthetic prokaryote, cyanobacteria. The molecular mechanism of this clock has been efficiently dissected using a luciferase reporter gene that reports the status of the clock. A circadian clock gene cluster, kaiABC, has been cloned via rhythm mutants of cyanobacterium, Synechococcus, and many clock mutations mapped to the three kai genes. Although kai genes do not share any homology with clock genes so far identified in eukaryotes, analysis of their expression suggests that a negative feedback control of kaiC expression by KaiC generates the circadian oscillation and that KaiA functions as a positive factor to sustain this oscillation. BioEssays 22:10-15, 2000.     

Modelling circadian rhythms of protein KaiA, KaiB and KaiC interactions in cyanobacteria

Cyanobacteria are the simplest organisms known that exhibit circadian rhythms. The mechanism of circadian rhythm generation in cyanobacteria is different from eukaryotes. Based on the recent experiments about the interaction of KaiA, KaiB, and KaiC proteins with the generation of circadian rhythms in vitro, we developed a mathematical model to describe post-translational oscillations and the possible chemical reactions involved in the circadian clock mechanism of cyanobacteria. In this model, a series of differential equations, with linear kinetics for binding of proteins, Michaelis - Menten kinetics for enzymatic processes and a term including an explicit delay for dissociation of the KaiA/KaiB/phospho-KaiC complex, are proposed describing the dynamics of the chemistry. It is demonstrated that the mathematical system can lead to circadian oscillation within a range of parameter values.     

The circadian clocks of plants and cyanobacteria

Classical research on the circadian rhythms of plants helped to demonstrate that all living organisms utilize circadian clocks to adapt their day–night cycles and that the clock is the basis for photoperiodic time measurements. Molecular models for the circadian oscillator have now been elucidated in Drosophila, Neurospora, mice and cyanobacteria. All share a similar feedback structure, but key proteins in each of the oscillators are different. A plant clock model has yet to be proposed, but clock mutants of Arabidopsis are expected to reveal key proteins in the mechanism. Here we discuss how a self-sustained oscillation is established in eukaryotic and prokaryotic models, and the polyphyletic evolution of these clock systems.     

Cyanobacterial circadian clockwork: roles of KaiA,KaiB and the kaiBC promoter in regulating KaiC

Using model strains in which we ectopically express the cyanobacterial clock protein KaiC in cells from which the clock genes kaiA, kaiB and/or kaiC are deleted, we found that some features of circadian clocks in eukaryotic organisms are conserved in the clocks of prokaryotic cyanobacteria, but others are not. One unexpected difference is that the circadian autoregulatory feedback loop in cyanobacteria does not require specific clock gene promoters as it does in eukaryotes, because a heterologous promoter can functionally replace the kaiBC promoter. On the other hand, a similarity between eukaryotic clock proteins and the cyanobacterial KaiC protein is that KaiC is phosphorylated in vivo. The other essential clock proteins KaiA and KaiB modulate the status of KaiC phosphorylation; KaiA inhibits KaiC dephosphorylation and KaiB antagonizes this action of KaiA. Based upon an analysis of clock mutants, we conclude that the circadian period in cyanobacteria is determined by the phosphorylation status of KaiC and also by the degradation rate of KaiC. These observations are integrated into a model proposing rhythmic changes in chromosomal status.     

Precise circadian clocks in prokaryotic cyanobacteria

Prokaryotic cyanobacteria express robust circadian (daily) rhythms under the control of a timing mechanism that is independent of the cell division cycle. This biological clock orchestrates global regulation of gene expression and controls the timing of cell division. Proteins that may be involved in input pathways have been identified. Mutational screening has identified three clock genes that are organized as a gene cluster. The structure of cyanobacterial clock proteins, their phosphorylation, and regulation is described. A new model for the core clockwork in cyanobacteria proposes that rhythmic changes in the status of the chromosome underlie the rhythms of gene expression. Mixed-strain experiments demonstrate that this timekeeper confers adaptive value when different strains compete against each other.     

蓝藻生物钟分子机理的研究进展

蓝藻是具有内源性生物钟的简单生物.虽然蓝藻生物钟具有跟真核生物同样的基础特征,但其相关基因和蛋白质与真核生物没有同源性.蓝藻生物钟的核心是kai基因簇及其编码的蛋白KaiA,KaiB和KaiC.这三种Kai蛋白相互作用调节KaiC的磷酸化状态,从而产生昼夜节律信息.KaiC的磷酸化循环是昼夜节律的起博器,调控包括kai基因在内的相关基因的节律性表达.组氨酸蛋白激酶的磷酸化传递可将环境信息输入和将节律信息输出生物钟核心.     

Circadian programming in cyanobacteria.

Prokaryotic cyanobacteria express robust circadian (daily) rhythms under the control of a timing mechanism that is independent of the cell division cycle. This biological clock orchestrates global regulation of gene expression. Competition experiments demonstrate that fitness is enhanced when the circadian period is consonant with the period of the environmental cycle. Mutational analyses have identified three clock genes in the organism, one of which is related to DNA recombinases and helicases. We propose a new model for the core 'clockwork' that implicates rhythmic changes in the status of the chromosome that underly the rhythms of gene expression.     

LdpA: a component of the circadian clock senses redox state of the cell

The endogenous 24-h (circadian) rhythms exhibited by the cyanobacterium Synechococcus elongatus PCC 7942 and other organisms are entrained by a variety of environmental factors. In cyanobacteria, the mechanism that transduces environmental input signals to the central oscillator of the clock is not known. An earlier study identified ldpA as a gene involved in light-dependent modulation of the circadian period, and a candidate member of a clock-entraining input pathway. Here, we report that the LdpA protein is sensitive to the redox state of the cell and exhibits electron paramagnetic resonance spectra consistent with the presence of two Fe4S4 clusters. Moreover, LdpA copurifies with proteins previously shown to be integral parts of the circadian mechanism. We also demonstrate that LdpA affects both the absolute level and light-dependent variation in abundance of CikA, a key input pathway component. The data suggest a novel input pathway to the circadian oscillator in which LdpA is a component of the clock protein complex that senses the redox state of a cell.     

生物钟机制研究进展

由生物体内源性生物钟所产生的昼夜节律是近年来生命科学的研究热点之一。几种模型生物（蓝细菌、脉孢菌、拟南芥、果蝇、小鼠）的生物钟相关基因相继被克隆和鉴定,为理解昼夜节律的分子机制奠定了基础。振荡器蛋白对其编码基因的负反馈调控可能是不同生物的生物运作普遍机制,在此基础上,不同生物有不尽相同的调控方式;隐色素可能是高等生物的共同生物钟光受体。