Analysis of a new circadian mutant

描述(申请人提供)：我们在叙利亚仓鼠中发现了一种新的自发昼夜节律突变，这使动物的运动节律性具有强大的表型。突变纯合子动物的自由奔跑周期约为28小时，比野生型动物每天的周期长4小时。这些突变大多是通过化学诱变在小鼠身上产生的，对于我们迄今所知的哺乳动物细胞自主昼夜节律性的分子机制是至关重要的，但我们的知识远未完成，很明显，其他未知基因肯定对其功能起到了作用。此外，由于仓鼠在行为和生理研究方面比小鼠有许多优势，这种新的突变为研究哺乳动物的生物钟机制提供了独特的机会。我们建议从遗传、行为和分子三个层面分析这一新的突变。基于初步数据，基因分析将检验我们的假设，即突变是一个单一的常染色体共显性等位基因。Menaker实验室的工作将集中在行为表型上，并将探索野生型、杂合子和纯合子的雌鼠对恒定黑暗、恒定光照、单一光脉冲、不同光暗比的光周期和不同周期的携带周期的反应，这些数据将解决突变基因在昼夜分子机制中的作用及其对生物体生理的影响。格林实验室将对这些突变体进行分子和生化分析，以确定这种突变是否影响核心细胞内振荡器机制或某些系统水平的方面，如细胞间耦合。突变的性质将通过对已知的昼夜节律基因进行测序，并通过分析它们的表达水平来解决。这也将为最终通过全基因组测序鉴定突变奠定重要基础。昼夜节律调节了许多正常的生理和行为，而昼夜节律的扰乱增加了对各种环境侮辱的易感性。对潜在机制的了解对于控制这些有害影响至关重要。 与公共健康相关：从大脑延伸到外围每个结构的昼夜节律振荡器网络已经进化，以维持自然环境中的内部时间秩序。然而，这个系统是有压力的，内部的时间秩序被人类对他们自己的“非自然”环境所做的许多改变打乱了。为了减轻昼夜节律紊乱对人类健康的有害影响，有必要从分子细节上了解完整的和被破坏的系统。

DESCRIPTION (provided by applicant): We have discovered a new spontaneous circadian mutation in Syrian hamsters, which confers a robust phenotype on the animals' locomotor rhythmicity. The free running period of animals homozygous for the mutation is approximately 28 hours, 4 hours per daily cycle longer than that of wild type animals. Such mutations, most produced in mice by chemical mutagenesis, have been critical in working out what we know thus far about the molecular mechanism that generates cell autonomous circadian rhythmicity in mammals, but our knowledge is far from complete and it is clear that additional unidentified genes must contribute to its function. Furthermore, because hamsters provide many advantages over mice for behavioral and physiological studies, this new mutation provides a unique opportunity for studies of the circadian clock mechanism in mammals. We propose to analyze this new mutation at 3 levels: genetic, behavioral and molecular. Genetic analysis will test our assumption, based on preliminary data, that the mutation is a single, autosomal co-dominant allele. The work in the Menaker laboratory will be focused on the behavioral phenotype and will explore the responses of wild type, heterozygous and homozygous littermates of both sexes to constant darkness, constant light, single light pulses, photoperiods with different light/dark ratis and entraining cycles with different periods, These data will address the role of the mutant gene in the circadian molecular mechanism as well as its impact on the organism's physiology. The Green lab will perform molecular and biochemical analysis of these mutants to determine whether this mutation affects the core intracellular oscillator mechanism or some system-level aspect, such as intercellular coupling. The nature of the mutation will be addressed by sequencing of the known circadian genes, and by analysis of their expression levels. This will also lay important groundwork for the eventual identification of the mutation by whole-genome sequencing. Circadian rhythms modulate much of normal physiology and behavior and circadian disruptions increase vulnerability to a variety of environmental insults. Knowledge of the underlying mechanism is essential to controlling these deleterious effects. PUBLIC HEALTH RELEVANCE: The network of circadian oscillators that stretches from the brain to every structure in periphery has evolved to maintain internal temporal order in the natural environment. However, this system is stressed and internal temporal order is disrupted by many of the changes that humans have made to their own "unnatural" environments. In order to mitigate the deleterious effects of circadian disruption on human health, it is necessary to understand the intact and the disrupted system in molecular detail.