Slit-Robo Signalling Establishes a Sphingosine-1-Phosphate Gradient to Polarise Fin Mesenchyme and Establish Fin Morphology

Abstract

Immigration of mesenchymal cells into the growing fin and limb buds drives distal outgrowth, with subsequent tensile forces between these cells essential for fin and limb morphogenesis. Morphogens derived from the apical domain of the fin, orientate limb mesenchyme cell polarity, migration, division and adhesion. The zebrafish mutant stomp displays defects in fin morphogenesis including blister formation and associated loss of orientation and adhesion of immigrating fin mesenchyme cells. Positional cloning of stomp identified a mutation in the gene encoding the axon guidance ligand, Slit3. We provide evidence that Slit ligands derived from immigrating mesenchyme act via Robo receptors at the Apical Ectodermal Ridge (AER) to promote release of sphingosine-1-phosphate (S1P). S1P subsequently diffuses back to the mesenchyme to promote their polarisation, orientation, positioning and adhesion to the interstitial matrix of the fin fold. We thus demonstrate coordination of the Slit-Robo and S1P signalling pathways in fin fold morphogenesis. Our work introduces a mechanism regulating the orientation, positioning and adhesion of its constituent cells.

Echolocation-like model of directed cell migration within growing tissues

“…Details of the experimental system can be found in Ref. (66). Given the very narrow width of the fin, cell migration within this system is effectively two-dimensional.…”

“…(C, C’) Normalised tracks of distal (blue) and proximal (red) cells from embryos (C) and from simulations (C’). (D, D’) Log-scaled jitter plots of pFAK intensity in embryos (D) (replotted from (66), Fig. EV5) and R signals in simulation (D’).…”

“…Genetic experiments by Mahabaleshwar et. al (66) suggest that this migration is mediated by feedback signals between the cell and the AER. Slit ligands are produced by the migrating fibroblasts, which then act via its canonical receptor Robo at the AER to induce the release of Sphingosine-1-Phosphate (S1P).…”

“…In Ref. (66) the levels of the adhesion adhesion molecule pFAK (70) were analyzed across the fin fold. Comparing this data (replotted in Fig.…”

“…Values 10µm 2 s −1 0.1s −1 0.3molm −2 s −1 , 0.3s −1 10 8 molm −3 0.33µmmin −1 movies and pFAK intensity measurements were obtained as described in Ref. (66). Selected cells were manually segmented and tracked on Fiji.…”