期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2022
卷号：119
期号：34
DOI：10.1073/pnas.2206066119
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Significance
Neuromodulation is essential for adaptive animal behaviors among other physiological processes. It is essential to reliably manipulate neuromodulator pathways to understand their functions in animal physiology. In this study, we generated a CRISPR-Cas9-based guide library to target every G-Protein Coupled Receptor (GPCR) in the
Drosophila genome and applied it to the well-studied clock neuron network. Notably, these GPCRs are highly enriched and differentially expressed in this small network, making it an ideal candidate to investigate their function. We cell-type specifically mutated GPCRs highly efficiently with no background gene editing detected. Applying this strategy to a specific node of the clock network revealed a role for dopamine in prolonging daytime sleep, suggesting network-specific functions of dopamine receptors in sleep-wake regulation.
The metronome-like circadian regulation of sleep timing must still adapt to an uncertain environment. Recent studies in
Drosophila indicate that neuromodulation not only plays a key role in clock neuron synchronization but also affects interactions between the clock network and brain sleep centers. We show here that the targets of neuromodulators, G Protein Coupled Receptors (GPCRs), are highly enriched in the fly brain circadian clock network. Single-cell sequencing indicates that they are not only enriched but also differentially expressed and contribute to clock neuron identity. We generated a comprehensive guide library to mutagenize individual GPCRs in specific neurons and verified the strategy by introducing a targeted sequencing approach. Combined with a behavioral screen, the mutagenesis strategy revealed a role of dopamine in sleep regulation by identifying two dopamine receptors and a clock neuron subpopulation that gate the timing of sleep.
