Andante/CKIIB in the Drosophila Circadian Clock

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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.