View Program - 63rd Annual Drosophila Research Conference

Precise wiring of neural circuits is essential for the execution of vital functions, from breathing to complex motor movements. This connectivity is established during development as the axons of newborn neurons extend through the embryonic environment, sensing both attractive and repulsive cues via guidance receptors. In the central nervous system (CNS), commissural neurons send their axons across the midline to connect to contralateral targets. This is essential for left-right coordination, yet the underlying mechanisms are not fully understood. In flies, the guidance receptor Frazzled (Fra), like its human ortholog Deleted in Colorectal Cancer (Dcc), promotes axon growth toward the midline. Despite its critical role in axon guidance, our understanding of Fra/Dcc signaling remains incomplete. We performed an affinity purification-mass spectrometry screen in fly embryonic neurons to identify novel Fra interactors. We selected three candidates, the Toll-like receptor family member Toll-7, the iRhom pseudoprotease family member rhomboid-5 (rho-5), and the exportin embargoed (emb), all expressed in the embryonic CNS, for further investigation. Toll-7 regulates neuronal connectivity in the fly olfactory system, but its involvement in Fra signaling has not been investigated. Human iRhoms promote ADAM17 metalloprotease activity, which was shown by our lab to cleave and positively regulate the functions of both Fra and Dcc, making rho-5 a promising candidate regulator of Fra signaling. Finally, as nuclear export antagonizes the transcriptional activity of Fra, emb may also modulate Fra signaling. Our goal is to characterize the roles of Toll-7, rho-5, and emb in Fra and Dcc signaling. We show that Toll-7 and emb biochemically interact with Fra in S2R+ cells. Importantly, we find that all three candidates modulate midline crossing: Toll-7 and rho-5 promote crossing, while emb antagonizes this process. Given their physical association with Fra, it is likely that the candidates regulate midline crossing via the Fra pathway. Future experiments will determine whether Toll-7, rho-5 and emb function in the Fra pathway and if these genes play conserved roles in vertebrate Dcc signaling. Our research will shed light on uncharacterized mechanisms of Fra/Dcc signaling and axon guidance in Drosophila and vertebrates. These findings will provide insight into the mechanisms driving assembly of neural circuits, which may inform interventions to promote neuronal repair.

Investigating mechanisms of Frazzled/Dcc signaling in axon guidance