Abstract
Circadian rhythms have long been established as a contributor to maintaining homeostasis in many physiological aspects of life. Much of the research on homeostasis and plasticity is done on a shorter time scale, attributed to neural activity. However, longer time scale shifts with morphological restructuring are also present in the circadian network. In the Drosophila brain, small ventral lateral neurons (s LNvs) have axon projections that extend towards dorsal neuron clusters. At the ends of these projections, axon spread cycles daily, defasciculating during the day and fasciculating at night. The mechanisms and effects of this phenomenon remains largely uncharacterized, but prior research has identified various transcription factors and GTPases that are associated with changes in structure. Single-cell seq shows a peak in robo3 RNA during the day and little production at night. This confirms circadian characteristics of robo3 at an RNA level and likely to translate to the protein level. Using various mutant strains, we demonstrate that robo3 is implicated in cellular restructuring processes as well. An adult-specific expression system showed that there is a sustained plasticity in adult flies, wherein inducing RNA knockdown or overexpression could still significantly change the morphology at the axon terminals. Robo3 knockdowns yield a defasciculated (open) structure, while robo3 overexpression causes more fasciculation (closed). In both cases, the degree of fasciculation stayed constant between the day and night. Additionally, we identify Opa as a potential upstream regulator of robo3. There is a conserved motif and co-expression in s-LNvs, hinting at a coordinated transcriptional network. Opa mutants also display similar effects to robo3 strains, further confirming its role in the circuitry governing these morphological changes. These findings provide insight into how robo3 and Opa fit into a larger picture of daily plasticity at a cellular level.