Casein kinase Iε Does Not Rescue double-time Function in Drosophila Despite Evolutionarily Conserved Roles in the Circadian Clock

Abstract

double-time (dbt) is a casein kinase gene involved in cell survival, proliferation, and circadian rhythms in the fruit fly, Drosophila melanogaster. Genetic and biochemical studies have shown that dbt and its mammalian ortholog casein kinase Iε (hckIε) regulate the circadian phosphorylation of period (per), thus controlling per subcellular localization and stability. Mutations in these kinases can shorten the circadian period in both mammals and Drosophila. Since similar activities in circadian clock have been described for these kinases, we investigated whether the expression of mammalian casein kinase I can replace the activity of dbt in flies. Global expression of the full-length dbt rescued lethality of the null mutant dbt ^revVIII and rescued flies showed normal locomotor activity rhythms. Global expression of dbt also restored the locomotor activity rhythm of the arrhythmic genotype, dbt^ar /dbt^revVIII . In contrast, global expression of hckIε or hckIα did not rescue lethality or locomotor activity of dbt mutants. Furthermore dbt overexpression in wild-type clock cells had only a small effect on period length, whereas hckIε expression in clock cells greatly lengthened period to ~30.5 hours and increased the number of arrhythmic flies. These results indicate that hckIε cannot replace the activity of dbt in flies despite the high degree of similarity in primary sequence and kinase function. Moreover, expression of hckIε in flies appears to interfere with dbt activity. Thus, caution should be used in interpreting assays that measure activity of mammalian casein kinase mutants in Drosophila, or that employ vertebrate CKI in studies of dPER phosphorylations.