Circadian Rhythms of Gene Expression in Cyanobacteria

所有生物复杂性水平的生物体都表现出由内源性生化振荡器控制的昼夜节律。许多生理和细胞过程，包括睡眠/清醒、体温、体内平衡功能、基因表达、细胞分裂和酶活性，都受这些“生物钟”的调节。此外，在植物和动物生殖过程的光周期计时中，这些振荡器也是测量日长的计时器。因此，了解生物钟的生化机制具有重要的生物学意义。这些生物钟的生化性质一直难以捉摸，但它们的显著特性（持久性、温度补偿和携带）在从细菌到哺乳动物的所有生物体内都是保守的，这些生物体内的昼夜节律行为已经被观察到。这表明，生物化学机制也是保守的——如果不是完全同源，那么至少就时钟的一般成分和组成而言。本提案中描述的揭示昼夜节律振荡器机制的方法侧重于最不复杂和最具技术可接近的生物，其中生物钟已被证明，蓝藻，聚球菌属菌株PCC 7942。这种生物的技术优势在于它的基因组小，易于遗传操作，并且已经开发出一种生物发光菌株，其中昼夜节律基因表达的监测是目前可用的任何系统中最灵活和最方便的。在一次实验中，筛选数千个菌落，以寻找破坏生物钟或导致异常节律行为的突变是可能的。因此，这个蓝藻系统有详细的分子/遗传分析和时钟调查极好的工具。本项目将使用该系统来解决生物节律性的三个方面：(1)识别和表征参与产生昼夜节律的基因，(2)表征振荡器控制节奏输出的机制，以及(3)评估生物体从其昼夜节律振荡器中获得的适应性优势。所有生物复杂程度的生物体都表现出由内部生化振荡器控制的昼夜节律。许多生理和细胞过程，包括睡眠/清醒、体温、体内平衡功能、基因表达、细胞分裂和酶活性，都受这些“生物钟”的调节。此外，这些时钟也是植物和动物生殖过程光周期计时中测量日长度的计时器。精神病学和医学研究表明，昼夜节律与某些形式的抑郁症、“时差反应”、药物耐受性/有效性、睡眠障碍和人类生理的其他方面有关。了解这些生物钟的机制一直是难以捉摸的，但一个已经出现的原则是，它们的主要特性在所有生物中都是保守的，在这些生物中，从细菌到哺乳动物的昼夜节律行为已经被观察到。在这个项目中描述的揭示昼夜节律振荡器机制的方法侧重于使用最不复杂和最具技术可接近的生物，其中生物钟已被证明（蓝藻），以发现时钟在分子水平上的工作原理。从这项工作中获得的关于生物钟机制的信息具有基本的生物学意义，并可能导致对农业以及心理健康和其他人类疾病的诊断和治疗有用的见解。* * * ? ?

Abstract Johnson 9633267 Organisms at all levels of biological complexity manifest circadian (daily) rhythms which are controlled by an endogenous biochemical oscillator. Many physiological and cellular processes, including sleeping/waking, body temperature, homeostatic functions, gene expression, cell division, and enzymatic activities, are regulated by these "biological clocks". In addition, these oscillators are also the timers that measure the daylength in photoperiodic timing of reproductive processes in plants and animals. Therefore, understanding the biochemical mechanism of circadian clocks is of fundamental biological interest . The biochemical nature of these biological clocks has been elusive, but their salient properties (persistence, temperature compensation, and entrainment) are conserved in all organisms in which circadian behavior has been observed, from bacteria to mammals. This suggests that the biochemical mechanism has also been conserved--if not in exact homology, then at least in terms of the general components and composition of the clock. The approach to unveiling the mechanism of circadian oscillators described in this proposal focuses on the least-complex and most technically-approachable organism in which a biological clock has been demonstrated, the cyanobacterium, Synechococcus sp. strain PCC 7942. The technical advantages of this organism are that it has a small genome which is easily manipulated genetically, and that a bioluminescent strain has been developed in which the monitoring of circadian gene expression is the most flexible and facile of any system presently available. Screening of thousands of colonies for mutations that disrupt the clock or otherwise cause aberrant rhythmic behavior is possible within a single experiment. Therefore, this cyanobacterial system has excellent tools for detailed molecular/genetic analyses and for clock investigations. This project will use this system to address three aspects of biological rhythmicity: (1) identi fication and characterization of genes that are involved in generating circadian rhythms, (2) characterization of the mechanism by which the oscillator controls rhythmic outputs, and (3) assessment of the fitness advantage that organisms derive from their circadian oscillators. %%% Organisms at all levels of biological complexity manifest circadian (daily) rhythms which are controlled by an internal biochemical oscillator. Many physiological and cellular processes, including sleeping/waking, body temperature, homeostatic functions, gene expression, cell division, and enzymatic activities, are regulated by these "biological clocks." In addition, these clocks are also the timers that measure the daylength in photoperiodic timing of reproductive processes in plants and animals. Psychiatric and medical studies have shown that circadian rhythms are involved in some forms of depressive illness, "jet lag," drug tolerance/efficacy, sleep disorders, and other aspects of human physiology. Understanding the mechanism of these biological clocks has been elusive, but one principle which has emerged is that their major properties are conserved in all organisms in which circadian behavior has been observed, from bacteria to mammals. The approach to unveiling the mechanism of circadian oscillators described in this project focuses on using the least-complex and most technically-approachable organism in which a biological clock has been demonstrated (a cyanobacterium) to discover how the clock works at a molecular level. Information gained from this work about the mechanism of circadian clocks is of fundamental biological interest and may lead to insights which will be useful in agriculture and in the diagnosis and treatment of mental health and other human disorders. *** ??