In a groundbreaking study published in Nature, researchers have uncovered a sophisticated molecular mechanism that ensures precise synaptic partner matching in the fruit fly olfactory circuit. This discovery centers around a family of evolutionarily conserved immunoglobulin ligand–receptor pairs—specifically Hibris (Hbs)/Sticks and stones (Sns) and Kin of irre (Kirre)—which orchestrate repulsive interactions that prevent erroneous neuronal connections. These findings not only challenge long-standing perceptions about the functions of these molecules but also provide vital insights into the fundamental principles of neural circuit assembly.
The olfactory system of Drosophila melanogaster serves as an exceptional model for studying synaptic specificity due to its well-characterized architecture and genetic tractability. Within this system, odorant receptor neurons (ORNs) project to distinct glomeruli where they synapse with projection neurons (PNs). Ensuring that ORNs connect with their correct PN partners is crucial for accurate olfactory perception. Previous research had suggested that the Hbs/Sns–Kirre ligand–receptor pairs primarily mediated attractive interactions during processes such as myoblast fusion and nephrocyte function. However, emerging evidence indicated a more nuanced role, possibly involving repulsion.
To unravel this complexity, the research team employed an array of genetic manipulation techniques, including mutants and targeted RNA interference (RNAi), to dissect the roles of Kirre, Hbs, and Sns in synaptic partner matching. Strikingly, loss of function in either kirre, hbs, or sns resulted in a consistent phenotype: VA1v ORN axons mistargeted to the neighboring VA1d glomerulus. These phenotypes were recapitulated when Kirre was specifically knocked down in VA1v-ORNs or Hbs/Sns knocked down in VA1d-PNs, clearly demonstrating that these molecules actively prevent inappropriate synaptic connections.
Beyond loss-of-function approaches, the team assessed the consequences of Kirre overexpression within VA1d-ORNs, whose corresponding PNs express high levels of Hbs and Sns. Overexpression led to significant mistargeting of ORN axons to inappropriate glomeruli, such as VA1v and DA1. This miswiring phenotype underscored Kirre’s cell-autonomous role as a repulsive receptor, acting to restrict axonal targeting. Intriguingly, when Kirre overexpression occurred in flies mutated for either hbs, sns, or both, the mistargeting phenotype was notably suppressed, reinforcing the concept that Hbs/Sns serve as repulsive ligands for Kirre.
The authors also explored whether homophilic interactions—whereby Kirre binds to itself across membranes—might contribute to synaptic specificity. However, experiments in kirre hemizygous mutants showed that Kirre overexpression-induced mistargeting persisted unabated, effectively ruling out a role for homophilic Kirre interactions during synaptic partner matching. This distinction was crucial, as it highlighted heterophilic repulsion as the guiding mechanism to ensure ORN axons avoid forming synapses with incorrect PNs.
The study further demonstrated that multiple cell surface protein (CSP) pairs operate combinatorially to orchestrate precise neural wiring. Besides Hbs/Sns–Kirre, the researchers identified Fili–Kek1 and Toll2–Ptp10D pairs that also mediate complementary repulsive interactions. Importantly, genetic interaction analyses revealed that these pathways function independently since suppressing sns and hbs expression did not mitigate phenotypes caused by Kek1 overexpression. This modular arrangement of CSP-mediated repulsion likely provides the olfactory circuit with robustness and flexibility in wiring specificity.
Extending their investigation into synaptic outcomes, the team employed Bruchpilot-Short, a marker for presynaptic active zones, to visualize synapse formation by mistargeted ORN axons. The presence of this marker within mistargeted projections strongly suggests that these aberrant axons attempt to establish functional synapses with inappropriate PN targets. This ectopic connectivity could have profound implications for neural circuit function and sensory processing.
While biochemical and structural studies had previously confirmed direct binding between Hbs/Sns and Kirre, the authors encountered a surprising result when examining the other CSP pairs. In vitro and tissue-based binding assays failed to detect direct interactions between Fili and Kek1 or between Toll2 and Ptp10D, leaving the biochemical basis of these repulsive mechanisms enigmatic. The authors proposed several hypotheses, including the involvement of unidentified co-factors, post-translational modifications, or specific physiological conditions that their assays were unable to replicate.
This work significantly advances our understanding of how precise neural circuits are sculpted by repulsive molecular codes. By revealing that repulsion—not only attraction—plays a decisive role in synaptic partner matching, it challenges prevailing paradigms and opens new avenues for studying the molecular logic of neuronal wiring. Moreover, the demonstration that multiple CSP ligand–receptor pairs function additively and independently hints at an intricate combinatorial code that endows the nervous system with exquisite specificity.
Beyond its immediate implications for olfactory wiring, these findings may bear relevance for broader neurodevelopmental processes and disorders associated with synaptic miswiring. Understanding the molecular cues that prevent inappropriate synaptic connections could ultimately inform therapeutic strategies aimed at restoring or modulating neural circuits in disease contexts. Given the evolutionary conservation of these molecules, similar repulsive mechanisms may operate in vertebrate systems, including humans.
The study’s integrative approach—melding genetic, molecular, and imaging techniques—exemplifies the power of multidisciplinary strategies in unraveling complex biological phenomena. It reinforces the notion that synaptic specificity arises from a delicate balance between attraction and repulsion, fine-tuned by an array of cell surface proteins acting in concert. This conceptual framework will undoubtedly inspire future research aimed at decoding the synaptic connectivity map across diverse neural systems.
As neural circuit assembly continues to captivate neuroscientists, findings such as these shine a spotlight on the sophisticated molecular dialogues that guide neurons to their precise partners. The discovery of heterogeneous repulsive interactions mediated by Hbs/Sns–Kirre and allied CSP pairs enriches our mechanistic understanding and sets the stage for exploring how perturbations in such interactions contribute to neurodevelopmental disorders marked by defective connectivity.
In summary, this transformative research reveals that repulsions mediated by specific immunoglobulin superfamily proteins are integral to instructing synaptic partner matching in the Drosophila olfactory circuit. The identification of distinct ligand–receptor pairs operating in parallel pathways to forestall miswiring underscores the complexity and elegance of molecular codes driving neural circuit formation. As the field moves forward, elucidating the biochemical nuances underlying these interactions promises to illuminate fundamental principles of brain wiring with far-reaching implications.
Subject of Research: Mechanisms of synaptic partner matching and neural circuit specificity in the Drosophila olfactory system.
Article Title: Repulsions instruct synaptic partner matching in an olfactory circuit.
Article References: Li, Z., Lyu, C., Xu, C. et al. Repulsions instruct synaptic partner matching in an olfactory circuit. Nature (2025). https://doi.org/10.1038/s41586-025-09768-4
Image Credits: AI Generated
