Andante/CKIIB in the Drosophila Circadian Clock

描述（由申请人提供）：了解行为的细胞和分子基础是当代神经科学研究的主要目标，在这方面，果蝇已被证明是行为和神经系统分子遗传学研究的优秀模型系统。这个应用程序提出了一种被称为行板的果蝇昼夜节律突变的研究。该突变体表现出昼夜节律长周期表型（比野生型长约2小时），这种表型在成虫运动活动和成虫羽化节律中也被观察到，这与对分子振荡器的影响一致。此外，行板的遗传镶嵌分析表明，在头部（可能是神经）组织中有功能需求。行板突变已被定位到X染色体10E区域内的一个小基因组区间。最近，我们发现了该基因的新的转座子插入等位基因，这使我们能够开始定义行板转录单元的物理极限。我们的初步研究强烈表明，Andante编码酪蛋白激酶II (CKII) β同源物，这是CKII α / β全酶的已知调控亚基。鉴于已知翻译后调节对时钟功能的重要性，我们假设行板产物（ckii β）通过ckii α亚基调节分子振荡器的一个或多个元件的磷酸化（以及活性或稳定性）。在本申请中，我们建议：(1)确定行板基因的分子界限并验证其身份；(2)通过对现有行板点突变体的特征分析，探索行板表型的分子基础；(3)确定哪些组织和细胞类型需要行板功能来维持正常的昼夜周期；(4)明确行板产物（即CKIIbeta）调节生物钟功能的生化机制；(5)通过创建和行为表征α亚基变体，直接评估ckii α活性的昼夜节律需求。由于生物钟机制在果蝇和哺乳动物之间基本上是保守的，我们在果蝇身上提出的研究对于理解人类生物钟和治疗由昼夜节律时间改变引起的人类病理生理状态具有明显的影响。

DESCRIPTION (provided by applicant): Understanding the cellular and molecular bases of behavior is a cardinal goal of contemporary neuroscience research, and in this regard, the fruit fly Drosophila has proven to be an excellent model system for molecular genetic investigations of behavior and the nervous system. This application proposes studies of a Drosophila circadian mutant known as Andante. This mutant exhibits a circadian long-period phenotype (about 2h longer than the wild type), and such a phenotype is observed for adult locomotor activity and adult eclosion rhythms, consistent with an effect on the molecular oscillator. In addition, a genetic mosaic analysis of Andante indicates a functional requirement within head (and presumably neural) tissues. The Andante mutation has been mapped to a small genomic interval within region 10E of the X chromosome. Recently, we identified new transposon-insertion alleles of the gene, and this has enabled us to begin to define the physical limits of the Andante transcription unit. Our preliminary studies strongly suggest that Andante encodes a Casein Kinase II (CKII) beta ortholog, a known regulatory subunit of the CKIIalpha/beta holoenzyme. Given the known importance of post-translational regulation for clock function, we hypothesize that Andante product (CKIIbeta) acts through the CKIIalpha subunit to regulate phosphorylation (and activity or stability) of one or more elements of the molecular oscillator. In this application, we propose to: (1) define the molecular limits and verify the identity of the Andante gene; (2) explore the molecular basis of the Andante phenotype by characterizing an existing Andante point mutant; (3) determine which tissues and cell types require Andante function for normal circadian periodicity; (4) define the biochemical mechanism through which Andante product (i.e., CKIIbeta) regulates clock function; and (5) directly assess the circadian requirement for CKIIalpha activity by creating and behaviorally characterizing alpha subunit variants. Because the clock mechanism is fundamentally conserved between flies and mammals, our proposed studies in Drosophila have obvious ramifications for an understanding of the human circadian clock and the treatment of human pathophysiological states that arise from alterations of circadian timing.