L1CAM/Neuroglian controls the axon-axon interactions establishing layered and lobular mushroom body architecture.
Bottom Line: We demonstrate that the Drosophila melanogaster L1CAM homologue Neuroglian mediates adhesion between functionally distinct mushroom body axon populations to enforce and control appropriate projections into distinct axonal layers and lobes essential for olfactory learning and memory.For functional cluster formation, intracellular Ankyrin2 association is sufficient on one side of the trans-axonal complex whereas Moesin association is likely required simultaneously in both interacting axonal populations.Together, our results provide novel mechanistic insights into cell adhesion molecule-mediated axon-axon interactions that enable precise assembly of complex neuronal circuits.
Affiliation: Friedrich Miescher Institute for Biomedical Research, 4058 Basel, Switzerland University of Basel, 4003 Basel, Switzerland.Show MeSH
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Mentions: We then analyzed the cell type–specific requirements of extracellular adhesion using the nrg849 mutation, which causes an S213L exchange within the second Ig domain. It was previously reported that this mutation completely abolished Nrg-dependent homophilic cell–cell interactions in a Drosophila S2 cell aggregation assay (Goossens et al., 2011). However, a complete loss of adhesive properties of the NrgS213L protein in vivo is not consistent with the observation that nrg849 mutant animals survived to adulthood while nrg14- and nrgΔIg3–4 mutants that completely lack the extracellular Ig domains 3 and 4 die as late embryos (Bieber et al., 1989; Enneking et al., 2013). Interestingly, the potential analogous human L1CAM disease mutation H210Q (Jouet et al., 1994; Vits et al., 1994) reduces homophilic L1-L1 adhesion but efficiently binds to wild-type L1 protein (Castellani et al., 2002). To address potential association between NrgS213L and wild-type Nrg, we performed S2 cell aggregation assays using transient transfection of fluorescently tagged proteins. While expression of Nrg lacking Ig domains 3/4 did not induce cell cluster formation, we observed efficient clustering of cells expressing GFP-tagged NrgS213L (Fig. S3). This indicates at least partial homophilic binding activity consistent with the hypomorphic nature of the nrg849 mutation in vivo. In addition, we observed efficient association between NrgS213L and wild-type Nrg180-expressing cells, demonstrating that mutant and wild-type proteins can form functional homophilic interactions (Fig. S3). Based on these results, we next tested in vivo whether formation of a trans-axonal complex between wild-type and mutant NrgS213L proteins may be sufficient to rescue MB development. Indeed, expression of wild-type Nrg only in α′β′ neurons efficiently restored pedunculus projections and formation of α′β′ lobes in nrg849 mutants (Fig. 7, A–C and E). However, later ingrowing mutant αβ neurons failed to use these wild-type Nrg180-expressing neurons as a template, as indicated by the absence of αβ lobes (Fig. 7, C′ and F). Consistent with wild-type Nrg180 being required in the ingrowing neuronal subtype, we did not observe any rescue of mutant α′β′ or αβ MB axonal projections when wild-type Nrg180 was expressed in γ neurons using two different Gal4 lines (Fig. 7, D, D′, E, and F). Together, these rescue experiments revealed striking differential requirements of the extra- and intracellular domains of Nrg. These differences were particularly evident when comparing the α′β′ rescues in the different mutant backgrounds. The presence of wild-type Nrg180 in α′β′ neurons efficiently restored α′β′ but not αβ projections in nrg849 mutants, whereas it was sufficient to restore projections of both α′β′ and αβ neurons in nrg14; P[nrg180_ΔFIGQY] mutant animals (Fig. S4).
Affiliation: Friedrich Miescher Institute for Biomedical Research, 4058 Basel, Switzerland University of Basel, 4003 Basel, Switzerland.