The MAP3Ks Dual Leucine Kinase (DLK) and Leucine Zipper Kinase (LZK) are essential mediators of axon damage responses, but their responses are varied, complex, and incompletely understood. To characterize their functions in axon injury, we generated zebrafish mutants of each gene, labeled motor neurons (MN) and touch-sensing neurons in live zebrafish, precisely cut their axons with a laser, and assessed the ability of mutant axons to regenerate. DLK and LZK were required redundantly and cell autonomously for axon regeneration in MNs, but not in larval Rohon-Beard (RB) or adult dorsal root ganglion (DRG) sensory neurons. Surprisingly, in dlk lzk double mutants, the spared branches of wounded RB axons grew excessively, suggesting that these kinases inhibit regenerative sprouting in damaged axons. Uninjured trigeminal sensory axons also grew excessively in mutants when neighboring neurons were ablated, indicating that these MAP3Ks are general inhibitors of sensory axon growth. These results demonstrate that zebrafish DLK and LZK promote diverse injury responses, depending on the neuronal cell identity and type of axonal injury.

Epithelial cell properties are determined by the polarized distribution of membrane lipids, the cytoskeleton, and adhesive junctions. Epithelia are often profusely innervated, but little work has addressed how neurites affect epithelial organization. We previously found that basal keratinocytes in the zebrafish epidermis enclose axons in ensheathment channels sealed by autotypic junctions. Here we characterized how axons remodel cell membranes, the cytoskeleton, and junctions in basal keratinocytes. At the apical surface of basal keratinocytes, axons organized lipid microdomains quantitatively enriched in reporters for PI(4,5)P2 and liquid-ordered (Lo) membranes. Lipid microdomains supported the formation of cadherin-enriched F-actin protrusions, which wrapped around axons, likely initiating ensheathment. In the absence of axons, cadherin-enriched microdomains formed on basal cells, but did not organize into contiguous domains. Instead, these isolated domains formed heterotypic junctions with periderm cells, a distinct epithelial cell type. Thus, axon endings dramatically remodel polarized epithelial components and regulate epidermal adhesion. [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text]

Epithelial cell properties are determined by the polarized distribution of membrane lipids, the cytoskeleton, and adhesive junctions. Epithelia are often profusely innervated, but little work has addressed how contact with neurites affects the polarized organization of epithelial components. In previous work, we found that basal keratinocytes in the larval zebrafish epidermis wrap around axons to enclose them in ensheathment channels sealed by autotypic cell junctions. In this study, we used live imaging to characterize how sensory axons remodel cell membranes, the actin cytoskeleton, and adhesive junctions in basal keratinocytes. At the apical surface of basal keratinocytes, axons promoted the formation of lipid microdomains quantitatively enriched in reporters for PI(4,5)P2 and liquid-ordered (Lo) membranes. Lipid microdomains supported the formation of cadherin-enriched F-actin protrusions, which wrapped around axons, likely initiating the formation of ensheathment channels. Lo reporters, but not reporters of liquid-disordered (Ld) membranes, became progressively enriched at axon-associated membrane domains as autotypic junctions matured at ensheathment channels. In the absence of axons, cadherin-enriched lipid microdomains still formed on basal cell membranes, but were not organized into the contiguous domains normally associated with axons. Instead, these isolated domains formed ectopic heterotypic junctions with overlying periderm cells, a distinct epithelial cell type in the epidermis. Thus, axons inhibit the formation of epithelial heterotypic junctions by recruiting cadherin-rich lipid microdomains to form autotypic junctions at ensheathment channels. These findings demonstrate that sensory nerve endings dramatically remodel polarized epithelial components and regulate the adhesive properties of the epidermis.

The MAP3Ks Dual Leucine Kinase (DLK) and Leucine Zipper Kinase (LZK) are essential mediators of axon damage responses, but their responses are varied, complex, and incompletely understood. To characterize their functions in axon injury, we generated zebrafish mutants of each gene, labeled motor neurons (MN) and touch-sensing neurons in live zebrafish, precisely cut their axons with a laser, and assessed the ability of mutant axons to regenerate. DLK and LZK were required redundantly and cell autonomously for axon regeneration in MNs, but not in larval Rohon-Beard (RB) or adult dorsal root ganglion (DRG) sensory neurons. Surprisingly, in dlk lzk double mutants, the spared branches of wounded RB axons grew excessively, suggesting that these kinases inhibit regenerative sprouting in damaged axons. Uninjured trigeminal sensory axons also grew excessively in mutants when neighboring neurons were ablated, indicating that these MAP3Ks are general inhibitors of sensory axon growth. These results demonstrate that zebrafish DLK and LZK promote diverse injury responses, depending on the neuronal cell identity and type of axonal injury.