Dual-topology of collagen XV and tenascin C acts in concert to guide and shape developing motor axons

During development, motor axons are guided towards their muscle target by various extrinsic cues including extracellular matrix (ECM) proteins those identities remain poorly documented. Using single-cell RNA-sequencing of differentiating slow muscle progenitors (SMP) in zebrafish, we charaterized the SMP as a major source of ECM proteins that were computationally predicted to form a basement membrane-like structure tailored for motor axon guidance. Multiple in vivo and in vitro approaches further revealed that motor axon shape and growth relies on the timely expression of the attractive cue Collagen XV-B (ColXV-B) that locally provides motor axons with a permissive soft microenvironment and separately organizes the repulsive cue Tenascin C into a unique functional dual topology. Bioprinted micropatterns mimicking their unique topology provide compelling evidence that it represents a sufficient condition to elicit directional motor axon growth. Our study provides the first evidence that ECM topology and stiffness critically influence motor axon navigation in vertebrates with potential applications in regenerative medicine for peripheral nerve injury.