Modular control of endothelial sheet migration
Philip Vitorino and Tobias Meyer
Genes & Dev. 2008. 22: 3268-3281 doi: 10.1101/gad.1725808
j’ai demandé là-haut comment on explique par des gradients la fermetures des blastulas où on fait des trous
Growth factor-induced migration of endothelial cell monolayers enables embryonic development, wound healing, and angiogenesis. Although collective migration is widespread and therapeutically relevant, the underlying mechanism by which cell monolayers respond to growth factor, sense directional signals, induce motility, and coordinate individual cell movements is only partially understood. Here we used RNAi to identify 100 regulatory proteins that enhance or suppress endothelial sheet migration into cell-free space. We measured multiple live-cell migration parameters for all siRNA perturbations and found that each targeted protein primarily regulates one of four functional outputs: cell motility, directed migration, cell–cell coordination, or cell density. We demonstrate that cell motility regulators drive random, growth factor-independent motility in the presence or absence of open space. In contrast, directed migration regulators selectively transduce growth factor signals to direct cells along the monolayer boundary toward open space. Lastly, we found that regulators of cell–cell coordination are growth factor-independent and reorient randomly migrating cells inside the sheet when boundary cells begin to migrate. Thus, cells transition from random to collective migration through a modular control system, whereby growth factor signals convert boundary cells into pioneers, while cells inside the monolayer reorient and follow pioneers through growth factor-independent migration and cell–cell coordination.
Molecular components that direct sheet migration into cell-free space
The existence of a directed migration control module (Fig. 3E) argues strongly against a simple diffusive model for sheet migration whereby cells move randomly into unobstructed space. Because FGFR1 was identified as a member of this module, we compared the effect of titrating FGF concentration on random sheet migration against directed migration into cell-free space. While the cell velocity within the sheet was nearly constant in the absence or presence of FGF (Fig. 4A), the same increase in FGF caused a 10-fold increase in sheet migration. To investigate how FGF directs cells during sheet migration, we tracked individual cells within a migrating sheet. As expected, in regions distal to the open edge, cells moved randomly with similar speeds in both serum-starved and FGF treated cells. Near the sheet boundary, however, FGF-treated cells persistently migrate toward the cell-free space (Fig. 4B). To quantify this behavior, we counted the fraction of time cells spend migrating toward the monolayer edge (plus or minus 45°) and plotted the average value as a function of cell dis- tance from the open edge (Fig. 4C). In this analysis, a value of 0.25 reflects a random orientation since a 90° window represents one-quarter of a circle. Addition of FGF caused a significant increase in directed movement throughout the cell monolayer that was maximal near the sheet boundary, dropping off gradually at positions further from the edge. In contrast, serum-starved cells showed weaker orientation overall but maintained some directed movements near the sheet boundary (Fig. 4C). Next, we tracked cells transfected with siRNA pools against four members of the directed migration module, namely FGFR1, FRS2, GAB1, and PTEN, and compared their capacity for directed migration relative to control cells. Consistent with their role in directed migration, the first three genes, which slow sheet migration, showed a reduced orientation toward the cell-free band, while PTEN, which enhances sheet migration, showed a more pronounced orientation into cell-free space (Fig. 4D).
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