Pathways of molecular clock communication across the Drosophila melanogaster brain
Introduction: Behaviour is regulated through a network of neurons and transcription programs in the brain. Circadian behaviour is an innate behaviour that lends itself to robust and reproducible measurement, making it an ideal model for understanding the mechanisms that regulate behaviour. Circadian clocks are evolutionarily conserved transcription feedback loops that regulate circadian behaviour and circadian physiology. However, how circadian clocks communicate with each other to regulate circadian behaviour is not fully explored, primarily due to a lack of suitable tools. We developed a method called LABL (Johnstone et al., 2022) to measure circadian clock oscillations in distinct neurons, in vivo, using Drosophila melanogaster as a model organism. This method bypasses the need to measure locomotion behaviour, which is the terminal output of the circadian clocks, and allows us instead to measure circadian clocks directly to determine how circadian clocks can communicate with each other. Here, we genetically modify the Drosophila eye to determine which circadian clocks in the brain circadian neuronal network are responsive to the eye clock. Our data suggest that preferred communication pathways exist across this singular network, contradicting previous assumptions. Methods: The Drosophila Gal4/UAS and the LexA/LexAop systems allow for anatomically and temporally restricted expression of different genes. We exploited these tools to eliminate the Drosophila eye clock and silenced various neurons while simultaneously using LABL to monitor different brain clocks. We compared changes in transcriptional oscillations to changes in Drosophila behaviour. Results: Our data suggests the eye clocks communicate with the so-called LNd neuronal cluster of the circadian neuronal network to regulate waking behaviour in flies, while leaving other clocks unaffected. Our preliminary data suggest that this regulation is mediated through the LNvs. We propose a new model of circadian behaviour regulation that includes a flexible network of preferred communication pathways that are employed as a function of environmental input. Conclusion: Our data suggest that the circadian neuronal network is not a homogeneous network of clocks and neurons, as previously assumed. We favour a new model in which information passed through the neuronal network is context dependent and can be differently altered with genetic mutation. Our data also show that mutation-caused disruption to transcriptional programs in one part of a neuronal network can have different effects in transcription programs in other parts of the brain. This suggests that a mutation in one part of the brain can reveal its deleterious effect in a different part of the brain. Our data highlight the complexity of understanding behaviour genes in the context of behavioural disorders in the developing brain. Our continued work will provide insight into how disrupted behaviour genes are linked to behavioural disorders in children, at the molecular level.