Formation of the Posterior Lateral Line system in zebrafish is pioneered by the posterior Lateral Line (pLL) primordium, a group of about 150 cells that forms near the ear. While leading cells in the pLL primordium have a relatively mesenchymal morphology, trailing cells are more epithelial; they have distinct apical basal polarity and they reorganize to sequentially form nascent neuromasts or protoneuromasts. The pLL primordium begins migration toward the tip of the tail at about 22 hours post fertilization (hpf). Proliferation adds to the growth of the primordium, nevertheless, as the primordium migrates, the length of the column of cells undergoing collective migration progressively shrinks as cells stop migrating are deposited from the trailing end: cells that were incorporated into protoneuromasts are deposited as neuromasts, while cells that were not, are deposited between neuromasts as interneuromast cells. Eventually, the primordium ends its migration a day later after depositing 5-6 neuromasts and by resolving into 2-3 terminal neuromasts. Establishment of polarized Wnt and FGF signaling systems coordinates morphogenesis and migration of the primordium: Wnt signaling dominates at the leading end and is thought to determine the relatively mesenchymal morphology of leading cells, while FGF signaling dominates in the trailing end. There, FGF determines reorganization of groups of trailing cells to form rosettes as they constrict at their apical ends. Furthermore, FGF signaling determines the specification of a central cell in each rosette as a sensory hair cell progenitor and it helps determine collective migration of the pLL primordium cells. Wnt signaling promotes its own activity and at the same time drives expression of fgf3 and fgf10. However, leading cells do not respond to these FGF ligands because Wnt signaling simultaneously promotes expression of intracellular inhibitors of the FGF receptor. Instead, the FGFs activate FGF receptors and initiate FGF signaling at the trailing end of the primordium, where Wnt signaling is weakest. There, FGF signaling determines expression of the diffusible Wnt antagonist Dkk1b, which counteracts Wnt signaling to help establish stable FGF responsive centers. Once established, the trailing FGF signaling system coordinates morphogenesis of nascent neuromasts by simultaneously promoting the reorganization of cells into epithelial rosettes and by initiating expression of factors that help specify a sensory hair cell progenitor at the center of each forming neuromast. Over time, the leading domain with active Wnt signaling shrinks closer to the leading edge and additional FGF signaling centers form sequentially in its wake, each associated with formation of additional protoneuromasts. EPIDERMAL CONFINEMENT IS ESSENTIAL FOR COLLECTIVE MIGRATION DETERMINED BY SUPERFICIAL MIGRATORY CELLS THAT FORM A SHEATH AROUND DEEPER CELLS OF THE ZEBRAFISH POSTERIOR LATERAL LINE PRIMORDIUM During embryonic development, cells must navigate through a complex three-dimensional environment robustly and reproducibly. The zebrafish posterior lateral line primordium (PLLp), a group of approximately 120 cells which migrates from the otic vesicle to the tip of the tail, spearheading the development of the lateral line sensory system, is an excellent model to study such collective migration in an in vivo context. This system migrates in a channel formed by the underlying horizontal myoseptum and somites, and the overlying skin. While cells in the leading part of the PLLp are flat and have a more mesenchymal morphology, cells in the trailing part progressively reorganize to form epithelial rosettes, called protoneuromasts. These epithelial cells extend basal cryptic lamellipodia in the direction of migration in response to both chemokine and FGF signals. In this study, we show that, in addition to these cryptic lamellipodia, the core epithelial cells are in fact surrounded by a population of motile cells which extend actin-rich migratory processes apposed to the overlying skin. These thin cells wrap around the protoneuromasts, forming a continuous sheath of cells around the apical and lateral surface of the PLLp. The processes extended by these cells are highly polarized in the direction of migration and this directionality, like that of the basal lamellipodia, is dependent on FGF signaling. Consistent with interactions of sheath cells with the overlying skin contributing to migration, removal of the skin stalls migration. However, this is accompanied by some surprising changes. There is a profound change in the morphology of the sheath cells, with directional superficial lamellipodia being replaced with the appearance of undirected blebs or ruffles. Furthermore, removal of the skin not only affects underlying lamellipodia, it simultaneously alters the morphology and behavior of the deeper basal cryptic lamellipodia, even though these cells do not directly contact the skin. Directional actin-rich protrusions on both the apical and basal surface and migration are completely and simultaneously restored upon regrowth of the skin over the PLLp. We suggest that this system utilizes a circumferential sheath of motile cells to allow the internal epithelial cells to migrate collectively in the confined space of the horizontal myopseptum and that elastic confinement provided by the overlying skin is essential for effective collective migratory behavior of primordium cells.. SIGNALING AND MECHANICS-BASED MODELS OF LATERAL LINE NEUROMAST FORMATION AND DEPOSITION The posterior Lateral Line primordium is a group of about a hundred cells that migrates under the skin from near the ear to the tip of the tail periodically forming and depositing neuromasts to pioneer formation of the zebrafish Lateral Line sensory system. We describe how local activation coupled with long-range inhibition, operating via both signaling and mechanical interactions, potentially contribute to periodic formation and deposition of neuromasts by the migrating primordium. Wnt and Fgf signaling locally inhibit each other in leading and trailing zones of the migrating primordium, respectively, while Fgfs secreted by leading cells activate their receptors at a distance to promote Fgf signaling in a trailing zone. We use agent-based modelling to show how these interactions can determine periodic formation of Fgf signaling centers in the wake of a progressively shrinking, initially broad, Wnt domain. Center-biased Fgf signaling centers generated by these interactions provide the context for specification of central cells as sensory hair cell progenitors via Notch mediated lateral inhibition and the periodic reorganization of cells to form epithelial rosettes in the developing neuromasts. Independent observations suggest that primordium cells form adhesive links with cells within a defined neighborhood and pull them closer. As cells get closer, more cells can be pulled in. This local aggregation is inhibited by traction forces associated with collective migration, and the balance influences the emergent number and size of self-organized epithelial rosettes. We show how agent-based models based on such mechanical interactions can also make effective predictions about the pattern of neuromast formation and deposition by the migrating primordium under a variety of experimental conditions.
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