van Battum, E. Y. | Search bioRxiv

bioRxiv2024

Diverse and Location-Specific Roles of PlexinA2, PlexinA4, and NCAM in Developing Hippocampal Mossy Fibers

Mossy fibers (MFs) originate from dentate granule cells and innervate area CA3 of the hippocampus. Upon entry of CA3, MFs partition into two prominent axon bundles, the suprapyramidal tract (SPT) and infrapyramidal tract (IPT) and form lamina specific synaptic contacts in the stratum lucidum (SL) and stratum oriens (SO), respectively. Here we employed new mouse lines to dissect the function of Sema6A and its receptors, PlexinA2 (PlxnA2) and PlxnA4, in developing MFs. In Sema6a-/- mice, MF partitioning into SPT and IPT bundles is incomplete and IPT axons in the SO are overextended, while the SPT correctly innervates the SL. Loss of neuronal Sema6a results in defective MF patterning and we show that this involves Sema6A reverse signaling. Plxna4 controls MF partitioning, SPT axon bundling and laminar targeting to the SL, as well as IPT length. Many of these defects are replicated in mice deficient for PlxnA4 GAP catalytic activity, underscoring the importance of this GAP domain. MFs are tightly fasciculated in Plxna2-/- mice and fail to separate into SPT and IPT bundles, and defects are significantly reduced in PlxnA2 GAP mutants, highlighting the involvement of GAP-independent signaling events. To further explore the molecular basis of aberrant axon fasciculation, we employed anti-PlxnA2 dependent proximity biotinylation and identified several PlxnA2-associated Ig-CAM family members. We observed a genetic interaction between Plxna2 and Ncam1, but not Plxna4 and Ncam1, for SPT and IPT formation and positioning in CA3. Together, our studies provide insights into the multifaceted and overlapping, yet distinct, functions of PlxnA family members in orchestrating specific guidance decisions in developing MFs.

developmental biology↗

bioRxiv2023

Generation and characterization of Sema6aΔcyt conditional knockout mice

The axon guidance molecule Semaphorin-6A (SEMA6A) plays a key role during nervous system development. SEMA6A, classically known as a ligand for Plexin-A2 and -A4, is a transmembrane protein that can also elicit signaling via its intracellular domain in vitro. However, the physiological relevance of this reverse signaling route is largely unknown. We generated a new transgenic mouse model, Sema6a{Delta}cytfl/fl, in which the cytosolic part of SEMA6A can be conditionally removed using Cre-recombination. Upon Sema6a{Delta}cyt mutation, SEMA6A can only act as a ligand and reverse signaling is perturbed. Germline deletion of SEMA6As intracellular part results in developmental defects in axon pathfinding and neuron migration that partially phenocopy defects observed in full Sema6a knockout mice. These defects include disorganization of the anterior commissure, piriform cortex, lateral olfactory tract, thalamocortical and corticospinal white matter tracts, and defected neuron migration in the neocortex and cerebellum. Intriguingly, the hippocampal malformation described in Sema6a full knockout mice was not reproduced, suggesting a specific role for SEMA6A forward signaling in hippocampal development. Our results indicate that the intracellular domain of SEMA6A is essential for proper axon targeting and neuron migration, and provide the first proof of a SEMA6A reverse signaling pathway in vivo.

developmental biology↗