Dominant-Negative CK2α Induces Potent Effects on Circadian Rhythmicity

Circadian clocks organize the precise timing of cellular and behavioral events. In Drosophila, circadian clocks consist of negative feedback loops in which the clock component PERIOD (PER) represses its own transcription. PER phosphorylation is a critical step in timing the onset and termination of this feedback. The protein kinase CK2 has been linked to circadian timing, but the importance of this contribution is unclear; it is not certain where and when CK2 acts to regulate circadian rhythms. To determine its temporal and spatial functions, a dominant negative mutant of the catalytic alpha subunit, CK2αTik, was targeted to circadian neurons. Behaviorally, CK2αTik induces severe period lengthening (∼33 h), greater than nearly all known circadian mutant alleles, and abolishes detectable free-running behavioral rhythmicity at high levels of expression. CK2αTik, when targeted to a subset of pacemaker neurons, generates period splitting, resulting in flies exhibiting both long and near 24-h periods. These behavioral effects are evident even when CK2αTik expression is induced only during adulthood, implicating an acute role for CK2α function in circadian rhythms. CK2αTik expression results in reduced PER phosphorylation, delayed nuclear entry, and dampened cycling with elevated trough levels of PER. Heightened trough levels of per transcript accompany increased protein levels, suggesting that CK2αTik disturbs negative feedback of PER on its own transcription. Taken together, these in vivo data implicate a central role of CK2α function in timing PER negative feedback in adult circadian neurons.
Author Summary The molecular mechanism that governs organization of physiology and behavior into 24-h rhythms is a conserved transcriptional feedback process that is strikingly similar across distinct phyla. Notably, cyclic phosphorylation of negative feedback regulators is critical to time molecular rhythms. Indeed, mutation of a putative phosphoacceptor site in the human PERIOD2 gene, a key negative regulator, is associated with Advanced Sleep Phase Syndrome. This study reveals a critical role for the protein kinase CK2 for setting the period of behavioral and molecular oscillations in Drosophila. Circadian phenotypes due to CK2 disruption are due to a direct requirement in adult circadian pacemakers. These findings further demonstrate that CK2 modification of the negative feedback regulator PERIOD alters its cyclical phosphorylation, protein abundance, nuclear translocation, and transcriptional repression activity. These studies place CK2 as a central kinase in circadian timing.