Ca2ϩ signals are known to be important regulators of neurite outgrowth and steering. Here we show that inhibiting Ca 2ϩ influx through stretch-activated channels using various compounds, including a highly specific peptide isolated from Grammostola spatulata spider venom (GsMTx4), strongly accelerates the rate of neurite extension on diverse substrata and within the intact spinal cord. Consistent with the presence of stretch-activated channels, we show that Ca 2ϩ influx is triggered by hypotonic solutions, which can be partially blocked by GsMTx4. Finally, chelating local, but not global, Ca 2ϩ signals prevents the acceleration that is normally produced by GsMTx4. Blocking Ca 2ϩ influx through other channel types has little or opposite effects, but release from intracellular stores is required for maximal acceleration. Together, our data suggest that Ca 2ϩ functions at distinct microdomains in growth cones, with influx through mechanosensitive channels acting to inhibit outgrowth in opposition to influx through other plasma membrane channels and release from stores.

Intracellular Ca2+ signals control the development and regeneration of spinal axons downstream of chemical guidance cues, but little is known about the roles of mechanical cues in axon guidance. Here we show that transient receptor potential canonical 1 (TRPC1) subunits assemble mechanosensitive (MS) channels on Xenopus neuronal growth cones that regulate the extension and direction of axon outgrowth on rigid, but not compliant, substrata. Reducing expression of TRPC1 by antisense morpholinos inhibits the effects of MS channel blockers on axon outgrowth and local Ca2+ transients. Ca2+ influx through MS TRPC1 activates the protease calpain, which cleaves the integrin adaptor protein talin to reduce Src-dependent axon outgrowth, likely through altered adhesion turnover. We found that talin accumulates at the tips of dynamic filopodia, which is lost upon cleavage of talin by active calpain. This pathway may also be important in axon guidance decisions since asymmetric inhibition of MS TRPC1 is sufficient to induce growth cone turning. Together our results suggest that Ca2+ influx through MS TRPC1 on filopodia activates calpain to control growth cone turning during development.

The neural crest is a population of multipotent cells that migrates extensively throughout vertebrate embryos to form diverse structures. Mice mutant for the de novo DNA methyltransferase DNMT3b exhibit defects in two neural crest derivatives, the craniofacial skeleton and cardiac ventricular septum, suggesting that DNMT3b activity is necessary for neural crest development. Nevertheless, the requirement for DNMT3b specifically in neural crest cells, as opposed to interacting cell types, has not been determined. Using a conditional DNMT3b allele crossed to the neural crest cre drivers Wnt1-cre and Sox10-cre, neural crest DNMT3b mutants were generated. In both neural crest-specific and fully DNMT3b-mutant embryos, cranial neural crest cells exhibited only subtle migration defects, with increased numbers of dispersed cells trailing organized streams in the head. In spite of this, the resulting cranial ganglia, craniofacial skeleton, and heart developed normally when neural crest cells lacked DNMT3b. This indicates that DNTM3b is not necessary in cranial neural crest cells for their development. We conclude that defects in neural crest derivatives in DNMT3b mutant mice reflect a requirement for DNMT3b in lineages such as the branchial arch mesendoderm or the cardiac mesoderm that interact with neural crest cells during formation of these structures.

Nuclear receptor–binding, SET-domain containing 3 (NSD3) is the first protein methyltransferase essential for early neural crest development. NSD3 is required for neural crest gene expression but not for H3K36 dimethylation of most neural crest genes. NSD3-related methyltransferase activity independently regulates neural crest migration.

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