The development of cell polarity is an essential prerequisite for tissue morphogenesis during embryogenesis, particularly in the development of epithelia. In addition, oriented cell division can have a powerful influence on tissue morphogenesis. Here we identify a novel mode of polarized cell division that generates pairs of neural progenitors with mirror-symmetric polarity in the developing zebrafish neural tube and has dramatic consequences for the organization of embryonic tissue. We show that during neural rod formation the polarity protein Pard3 is localized to the cleavage furrow of dividing progenitors, and then mirror-symmetrically inherited by the two daughter cells. This allows the daughter cells to integrate into opposite sides of the developing neural tube. Furthermore, these mirror-symmetric divisions have powerful morphogenetic influence: when forced to occur in ectopic locations during neurulation, they orchestrate the development of mirror-image pattern formation and the consequent generation of ectopic neural tubes.

Dorsolateral bending of the neural plate, an undifferentiated pseudostratified epithelium, is essential for neural tube closure in the mouse spinal region. If dorsolateral bending fails, spina bifida results. In the present study, we investigated the molecular signals that regulate the formation of dorsolateral hinge points (DLHPs). We show that Bmp2 expression correlates with upper spinal neurulation (in which DLHPs are absent); that Bmp2-null embryos exhibit premature, exaggerated DLHPs; and that the local release of Bmp2 inhibits neural fold bending. Therefore, Bmp signalling is necessary and sufficient to inhibit DLHPs. By contrast, the Bmp antagonist noggin is expressed dorsally in neural folds containing DLHPs, noggin-null embryos show markedly reduced dorsolateral bending and local release of noggin stimulates bending. Hence, Bmp antagonism is both necessary and sufficient to induce dorsolateral bending. The local release of Shh suppresses dorsal noggin expression, explaining the absence of DLHPs at high spinal levels, where notochordal expression of Shh is strong. DLHPs 'break through' at low spinal levels, where Shh expression is weaker. Zic2 mutant embryos fail to express Bmp antagonists dorsally and lack DLHPs, developing severe spina bifida. Our findings reveal a molecular mechanism based on antagonism of Bmp signalling that underlies the regulation of DLHP formation during mouse spinal neural tube closure.

By analysing the cellular and subcellular events that occur in the centre of the developing zebrafish neural rod, we have uncovered a novel mechanism of cell polarisation during lumen formation. Cells from each side of the neural rod interdigitate across the tissue midline. This is necessary for localisation of apical junctional proteins to the region where cells intersect the tissue midline. Cells assemble a mirror-symmetric microtubule cytoskeleton around the tissue midline, which is necessary for the trafficking of proteins required for normal lumen formation, such as partitioning defective 3 and Rab11a to this point. This occurs in advance and is independent of the midline cell division that has been shown to have a powerful role in lumen organisation. To our knowledge, this is the first example of the initiation of apical polarisation part way along the length of a cell, rather than at a cell extremity. Although the midline division is not necessary for apical polarisation, it confers a morphogenetic advantage by efficiently eliminating cellular processes that would otherwise bridge the developing lumen.

ATP-gated P2X 2 channels undergo permeability changes through a process that is incompletely understood. In the present study, we used fluorescence resonance energy transfer (FRET) and electrophysiology to measure cytosolic gating motions in P2X 2 channels as they enter a state with increased permeability. P2X 2 channels underwent permeability changes with a time course that was similar to decreases in FRET between cyan fluorescent protein and yellow fluorescent protein attached to the cytosolic domain of P2X 2 channels. Wild-type and mutant channels that did not undergo permeability changes also did not show evidence of cytosolic gating motions. Moreover, immobilizing the cytosolic domain by tethering it to the plasma membrane prevented the switch in permeability and impaired the cytosolic gating motions. Both of these phenotypes were restored when the immobilizing tether was cleaved. The data provide a time-resolved measure of state-specific gating motions and suggest how a cytosolic domain may control ion channel permeability.

BackgroundMorphogenesis of the zebrafish neural tube requires the coordinated movement of many cells in both time and space. A good example of this is the movement of the cells in the zebrafish neural plate as they converge towards the dorsal midline before internalizing to form a neural keel. How these cells are regulated to ensure that they move together as a coherent tissue is unknown. Previous work in other systems has suggested that the underlying mesoderm may play a role in this process but this has not been shown directly in vivo.ResultsHere we analyze the roles of subjacent mesoderm in the coordination of neural cell movements during convergence of the zebrafish neural plate and neural keel formation. Live imaging demonstrates that the normal highly coordinated movements of neural plate cells are lost in the absence of underlying mesoderm and the movements of internalization and neural tube formation are severely disrupted. Despite this, neuroepithelial polarity develops in the abnormal neural primordium but the resulting tissue architecture is very disorganized.ConclusionsWe show that the movements of cells in the zebrafish neural plate are highly coordinated during the convergence and internalization movements of neurulation. Our results demonstrate that the underlying mesoderm is required for these coordinated cell movements in the zebrafish neural plate in vivo.