Abstract
The circadian clock in eukaryotes keeps time via transcriptional feedback loops. In the transcriptional feedback loop of animals, CLOCK activator complexes drive expression of PER repressor complex components which feedback to inhibit CLOCK activation until PER complexes are degraded, thus initiating the next round of CLOCK activation ~24 h later. Recently, we showed that a region of monarch CLOCK (CLK) analogous to that encoded by mammalian CLOCK exon 19 (CLKe19r) and the methyltransferase TRITHORAX (TRX) are required for CLK activation, PER-CLK binding, and PER repression and that TRX-dependent methylation of Heat Shock Protein 68 (HSP68) at arginine 45 (R45) is necessary for PER-CLK binding and PER repression. Given that CLK activation and PER repression complexes in Drosophila are comprised of different core components than in monarchs, we tested whether similar mechanisms are used for CLK activation and PER repression in Drosophila. We found that the CLKe19r, TRX and HSP68 are all required for CLK activation yet only HSP68, but not HSP68 R45 methylation, is required for PER repression in Drosophila. These results reveal a well-conserved CLK activation mechanism and a PER repression mechanism that retains HSP68 function but does not require TRX-dependent methylation.