PDF Signaling Is an Integral Part of the Drosophila Circadian Molecular Oscillator

Circadian clocks generate 24-hr rhythms in physiology and behavior. Despite numerous studies, it is still uncertain how circadian rhythms emerge from their molecular and neural constituents. Here, we demonstrate a tight connection between the molecular and neuronal circadian networks. Using fluorescent transcriptional reporters in a Drosophila ex vivo brain culture system, we identified a reciprocal negative regulation between the master circadian regulator CLK and expression of pdf, the main circadian neuropeptide. We show that PDF feedback is required for maintaining normal oscillation pattern in CLK-driven transcription. Interestingly, we found that CLK and neuronal firing suppresses pdf transcription, likely through a common pathway involving the transcription factors DHR38 and SR, establishing a direct link between electric activity and the circadian system. In sum, our work provides evidence for the existence of an uncharacterized CLK-PDF feedback loop that tightly wraps together the molecular oscillator with the circadian neuronal network in Drosophila.