Tyrosine phosphorylation of β-catenin, a component of adhesion complexes and of the Wnt pathway, affects cell adhesion, migration and gene transcription. By reducing β-catenin availability using shRNA-mediated gene silencing or expression of intracellular N-cadherin, we show that β-catenin is required for axon growth downstream of brain-derived neurotrophic factor (BDNF) signalling and hepatocyte growth factor (HGF) signalling. We demonstrate that the receptor tyrosine kinases (RTKs) Trk and Met interact with and phosphorylate β-catenin. Stimulation of Trk receptors by neurotrophins (NTs) results in phosphorylation of β-catenin at residue Y654, and increased axon growth and branching. Conversely, pharmacological inhibition of Trk or expression of a Y654F mutant blocks these effects. β-catenin phosphorylated at Y654 colocalizes with the cytoskeleton at growth cones. However, HGF, which also increases axon growth and branching, induces β-catenin phosphorylation at Y142 and a nuclear localization. Interestingly, dominant-negative ΔN-TCF4 abolishes the effects of HGF in axon growth and branching, but not that of NTs. We conclude that NT- and HGF-signalling differentially phosphorylate β-catenin, targeting this protein to distinct compartments to regulate axon morphogenesis by TCF4-transcription-dependent and -independent mechanisms. These results place β-catenin downstream of growth-factor–RTK signalling in axon differentiation.

Wnt factors regulate neural stem cell development and neuronal connectivity. Here we investigated whether Wnt-3a and Wnt-3, expressed in the developing spinal cord, regulate proliferation and the neuronal differentiation of spinal cord neural precursors (SCNP). Wnt-3a promoted a sustained increase of SCNP proliferation and decreased the expression of cyclin-dependent kinase inhibitors. In contrast, Wnt-3 transiently enhanced SCNP proliferation and increased neurogenesis through β-catenin signaling. Furthermore, both Wnt-3a and Wnt-3 stimulated neurite outgrowth in SCNP-derived neurons through β-catenin- and TCF4-dependent transcription. Glycogen synthase kinase-3β inhibitors mimicked Wnt signaling and promoted neurite outgrowth in established cultures. We conclude that Wnt-3a and Wnt-3 factors signal through the canonical Wnt/β-catenin pathway to regulate different aspects of SCNP development. These findings may be of therapeutic interest for the treatment of neurodegenerative diseases and nerve injury.

Axon morphogenesis is a complex process regulated by a variety of secreted molecules, including morphogens and growth factors, resulting in the establishment of the neuronal circuitry. Our previous work demonstrated that growth factors [Neurotrophins (NT) and Hepatocyte Growth Factor (HGF)] signal through β-catenin during axon morphogenesis. HGF signaling promotes axon outgrowth and branching by inducing β-catenin phosphorylation at Y142 and transcriptional regulation of T-Cell Factor (TCF) target genes. Here, we asked which genes are regulated by HGF signaling during axon morphogenesis. An array screening indicated that HGF signaling elevates the expression of chemokines of the CC and CXC families. In line with this, CCL7, CCL20, and CXCL2 significantly increase axon outgrowth in hippocampal neurons. Experiments using blocking antibodies and chemokine receptor antagonists demonstrate that chemokines act downstream of HGF signaling during axon morphogenesis. In addition, qPCR data demonstrates that CXCL2 and CCL5 expression is stimulated by HGF through Met/b-catenin/TCF pathway. These results identify CC family members and CXCL2 chemokines as novel regulators of axon morphogenesis downstream of HGF signaling.