Capillarity and active cell movement at mesendoderm translocation in the Xenopus gastrula.

During Xenopus gastrulation, leading edge mesendoderm (LEM) advances animally as a wedge-shaped cell mass over the vegetally moving blastocoel roof (BCR). We show that close contact across the BCR-LEM interface correlates with attenuated net advance of the LEM, which is pulled forward by tip cells while the remaining LEM frequently separates from the BCR. Nevertheless, lamellipodia persist on the detached LEM surface. They attach to adjacent LEM cells and depend on PDGF-A, cell-surface fibronectin and cadherin. We argue that active cell motility on the LEM surface prevents adverse capillary effects in the liquid LEM tissue as it moves by being pulled. It counters tissue surface-tension effects with oriented cell movement and bulges the LEM surface out to keep it close to the curved BCR without attaching to it. Proximity to the BCR is necessary, in turn, for the maintenance and orientation of lamellipodia that permit mass cell movement with minimal substratum contact. Together with a similar process in epithelial invagination, vertical telescoping, the cell movement at the LEM surface defines a novel type of cell rearrangement: vertical shearing.

Pdgf signalling guides neural crest contribution to the haematopoietic stem cell specification niche.

Haematopoietic stem cells (HSCs) support maintenance of the haematopoietic and immune systems throughout the life of vertebrates, and are the therapeutic component of bone marrow transplants. Understanding native specification of HSCs, to uncover key signals that might help improve in vitro directed differentiation protocols, has been a long-standing biomedical goal. The current impossibility of specifying true HSCs in vitro suggests that key signals remain unknown. We speculated that such signals might be presented by surrounding 'niche' cells, but no such cells have been defined. Here we demonstrate in zebrafish, that trunk neural crest (NC) physically associate with HSC precursors in the dorsal aorta (DA) just prior to initiation of the definitive haematopoietic program. Preventing association of the NC with the DA leads to loss of HSCs. Our results define NC as key cellular components of the HSC specification niche that can be profiled to identify unknown HSC specification signals.

PAPC mediates self/non-self-distinction during Snail1-dependent tissue separation.

Cleft-like boundaries represent a type of cell sorting boundary characterized by the presence of a physical gap between tissues. We studied the cleft-like ectoderm-mesoderm boundary in Xenopus laevis and zebrafish gastrulae. We identified the transcription factor Snail1 as being essential for tissue separation, showed that its expression in the mesoderm depends on noncanonical Wnt signaling, and demonstrated that it enables paraxial protocadherin (PAPC) to promote tissue separation through two novel functions. First, PAPC attenuates planar cell polarity signaling at the ectoderm-mesoderm boundary to lower cell adhesion and facilitate cleft formation. Second, PAPC controls formation of a distinct type of adhesive contact between mesoderm and ectoderm cells that shows properties of a cleft-like boundary at the single-cell level. It consists of short stretches of adherens junction-like contacts inserted between intermediate-sized contacts and large intercellular gaps. These roles of PAPC constitute a self/non-self-recognition mechanism that determines the site of boundary formation at the interface between PAPC-expressing and -nonexpressing cells.

Tissue cohesion and the mechanics of cell rearrangement.

Morphogenetic processes often involve the rapid rearrangement of cells held together by mutual adhesion. The dynamic nature of this adhesion endows tissues with liquid-like properties, such that large-scale shape changes appear as tissue flows. Generally, the resistance to flow (tissue viscosity) is expected to depend on the cohesion of a tissue (how strongly its cells adhere to each other), but the exact relationship between these parameters is not known. Here, we analyse the link between cohesion and viscosity to uncover basic mechanical principles of cell rearrangement. We show that for vertebrate and invertebrate tissues, viscosity varies in proportion to cohesion over a 200-fold range of values. We demonstrate that this proportionality is predicted by a cell-based model of tissue viscosity. To do so, we analyse cell adhesion in Xenopus embryonic tissues and determine a number of parameters, including tissue surface tension (as a measure of cohesion), cell contact fluctuation and cortical tension. In the tissues studied, the ratio of surface tension to viscosity, which has the dimension of a velocity, is 1.8 µm/min. This characteristic velocity reflects the rate of cell-cell boundary contraction during rearrangement, and sets a limit to rearrangement rates. Moreover, we propose that, in these tissues, cell movement is maximally efficient. Our approach to cell rearrangement mechanics links adhesion to the resistance of a tissue to plastic deformation, identifies the characteristic velocity of the process, and provides a basis for the comparison of tissues with mechanical properties that may vary by orders of magnitude.

EphA4-dependent Brachyury expression is required for dorsal mesoderm involution in the Xenopus gastrula.

Xenopus provides a well-studied model of vertebrate gastrulation, but a central feature, the movement of the mesoderm to the interior of the embryo, has received little attention. Here, we analyze mesoderm involution at the Xenopus dorsal blastopore lip. We show that a phase of rapid involution - peak involution - is intimately linked to an early stage of convergent extension, which involves differential cell migration in the prechordal mesoderm and a new movement of the chordamesoderm, radial convergence. The latter process depends on Xenopus Brachyury, the expression of which at the time of peak involution is controlled by signaling through the ephrin receptor, EphA4, its ligand ephrinB2 and its downstream effector p21-activated kinase. Our findings support a conserved role for Brachyury in blastopore morphogenesis.

Cadherin-dependent differential cell adhesion in Xenopus causes cell sorting in vitro but not in the embryo.

Adhesion differences between cell populations are in principle a source of strong morphogenetic forces promoting cell sorting, boundary formation and tissue positioning, and cadherins are main mediators of cell adhesion. However, a direct link between cadherin expression, differential adhesion and morphogenesis has not yet been determined for a specific process in vivo. To identify such a connection, we modulated the expression of C-cadherin in the Xenopus laevis gastrula, and combined this with direct measurements of cell adhesion-related parameters. Our results show that gastrulation is surprisingly tolerant of overall changes in adhesion. Also, as expected, experimentally generated, cadherin-based adhesion differences promote cell sorting in vitro. Importantly, however, such differences do not lead to the sorting of cells in the embryo, showing that differential adhesion is not sufficient to drive morphogenesis in this system. Compensatory recruitment of cadherin protein to contacts between cadherin-deprived and -overexpressing cells could contribute to the prevention of sorting in vivo.

Internalizing the vegetal cell mass before and during amphibian gastrulation: vegetal rotation and related movements.

The movement of the prospective mesoderm and endoderm to the interior of the amphibian embryo starts in the vegetal cell mass well before the onset of overt gastrulation. By an animally directed movement of cells, the vegetal mass constricts its outer part and expands its inner region including the blastocoel floor, in a process of pregastrulation emboly. Further internalization of the vegetal region has been studied in the Xenopus embryo. At the onset of gastrulation, vegetal rotation sets in at the periphery of the vegetal cell mass, first dorsally and then spreading laterally and ventrally. It consists of an intense inward surging of cells due to active cell rearrangements that can be observed in explants of the vegetal cell mass. In its course, the blastocoel floor expands further and becomes apposed to the blastocoel roof. The boundary between apposed floor and roof forms Brachet's cleft. Another effect of vegetal rotation is the downward and inward movement of the mesodermal marginal zone, constituting the first phase of involution. Together, the upward and outward movement of the peripheral vegetal mass and the downward and inward translocation of the marginal zone lead to an apparent rotation of the whole peripheral region of the gastrula. Vegetal rotation continues to contribute to endoderm internalization to near the end of gastrulation.

PDGF-A controls mesoderm cell orientation and radial intercalation during Xenopus gastrulation.

Radial intercalation is a common, yet poorly understood, morphogenetic process in the developing embryo. By analyzing cell rearrangement in the prechordal mesoderm during Xenopus gastrulation, we have identified a mechanism for radial intercalation. It involves cell orientation in response to a long-range signal mediated by platelet-derived growth factor (PDGF-A) and directional intercellular migration. When PDGF-A signaling is inhibited, prechordal mesoderm cells fail to orient towards the ectoderm, the endogenous source of PDGF-A, and no longer migrate towards it. Consequently, the prechordal mesoderm fails to spread during gastrulation. Orientation and directional migration can be rescued specifically by the expression of a short splicing isoform of PDGF-A, but not by a long matrix-binding isoform, consistent with a requirement for long-range signaling.