Real time, in vivo measurement of neuronal and peripheral clocks in Drosophila melanogaster

Abstract

Circadian clocks are highly conserved transcriptional regulators that control ~24-hour oscillations in gene expression, physiological function, and behavior. Circadian clocks exist in almost every tissue and are thought to control tissue-specific gene expression and function, synchronized by the brain clock. Many disease states are associated with loss of circadian regulation. How and when circadian clocks fail during pathogenesis remains largely unknown because it is currently difficult to monitor tissue-specific clock function in intact organisms. Here, we developed a method to directly measure the transcriptional oscillation of distinct neuronal and peripheral clocks in live, intact Drosophila, which we term Locally Activatable BioLuminescence, or LABL. Using this method, we observed that specific neuronal and peripheral clocks exhibit distinct transcriptional properties. Loss of the receptor for PDF, a circadian neurotransmitter critical for the function of the brain clock, disrupts circadian locomotor activity but not all tissue-specific circadian clocks. We found that, while peripheral clocks in non-neuronal tissues were less stable after the loss of PDF signaling, they continued to oscillate. We also demonstrate that distinct clocks exhibit differences in their loss of oscillatory amplitude or their change in period, depending on their anatomical location, mutation, or fly age. Our results demonstrate that LABL is an effective tool that allows rapid, affordable, and direct real-time monitoring of individual clocks in vivo.

Synaptic connectome of theDrosophilacircadian clock

“…However, the molecular clock architecture as well as the neuronal network motifs are highly conserved from humans to insects (Panda et al ., 2002, Helfrich-Förster, 2004). Hence, Drosophila melanogaster with its powerful genetic toolkit and a complete brain connectome represents an ideal system to decipher the clock network and its input and output pathways (Zheng et al ., 2018, Delventhal et al ., 2019, Johnstone et al ., 2022, Dorkenwald et al ., 2023, Schlegel et al ., 2023). Not surprisingly, the fly circadian clock, comprised of approximately 150 neurons, is extremely well characterized (Dubowy and Sehgal, 2017).…”

Cnidarians are CLOCKing in

“…This would not be so different from Drosophila : the oscillations in many peripheral tissues quickly decrease in amplitude upon transition to constant darkness, but in circadian neurons they remain robustly rhythmic in order to drive rhythmic behavior ( Johnstone et al, 2022 ; Stanewsky et al, 1997 ). Alternatively, the N. vectensis clock might rely heavily on post-transcriptional regulation.…”