Sleep is an evolutionarily conserved behavior, whose function is unknown. Here, we present a method for deep phenotyping of sleep in Drosophila, consisting of a high-resolution video imaging system, coupled with closed-loop laser perturbation to measure arousal threshold. To quantify sleep-associated microbehaviors, we trained a deep-learning network to annotate body parts in freely moving flies and developed a semi-supervised computational pipeline to classify behaviors. Quiescent flies exhibit a rich repertoire of microbehaviors, including proboscis pumping (PP) and haltere switches, which vary dynamically across the night. Using this system, we characterized the effects of optogenetically activating two putative sleep circuits. These data reveal that activating dFB neurons produces micromovements, inconsistent with sleep, while activating R5 neurons triggers PP followed by behavioral quiescence. Our findings suggest that sleep in Drosophila is polyphasic with different stages and set the stage for a rigorous analysis of sleep and other behaviors in this species.

Chromatin conformation reorganization is emerging as an important layer of regulation for gene expression and lineage specification. Yet, how lineage-specific transcription factors contribute to the establishment of cell type-specific 3D chromatin architecture in the immune cells remains unclear, especially for the late stages of T cell subset differentiation and maturation. Regulatory T cells (Treg) are mainly generated in the thymus as a subpopulation of T cells specializing in suppressing excessive immune responses. Here, by comprehensively mapping 3D chromatin organization during Treg cell differentiation, we show that Treg-specific chromatin structures were progressively established during its lineage specification, and highly associated with Treg signature gene expression. Additionally, the binding sites of Foxp3, a Treg lineage specifying transcription factor, were highly enriched at Treg-specific chromatin loop anchors. Further comparison of the chromatin interactions between wide-type Tregs versus Treg cells from Foxp3 knock-in/knockout or newly-generated Foxp3 domain-swap mutant mouse revealed that Foxp3 was essential for the establishment of Treg-specific 3D chromatin architecture, although it was not dependent on the formation of the Foxp3 domain-swapped dimer. These results highlighted an underappreciated role of Foxp3 in modulating Treg-specific 3D chromatin structure formation.

Natriuretic peptide system (NPS) and renin angiotensin aldosterone system (RAAS) function oppositely at multiple levels. While it has long been suspected that angiotensin II (ANGII) may directly suppress NPS activity, no clear evidence to date support this notion.