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It is widely accepted that tissue differentiation and morphogenesis in multicellular organisms are regulated by tightly controlled concentration gradients of morphogens. How exactly these gradients are formed, however, is much debated.
Here we present a study in living zebrafish embryos, where we explore the mechanism of concentration gradient formation of fibroblast growth factor 8 (Fgf8). Local concentrations and mobility parameters are determined with high precision by fluorescence correlation spectroscopy, combining different approaches of static-volume measurements of the ligand Fgf8 in extracellular space and scanning FCS measurements of its receptors in the cell membrane.
We show that Fgf8 molecules quickly spread as single molecules away from a source into the developing tissue by free Brownian diffusion. Upon binding, with similar affinities to Fgfr1 and Fgfr4, the ligand is taken up into the cells and hence cleared from the extracellular space. The equilibrium between a local morphogen production and the sink function of the receiving cells establishes the stable morphogen gradient. By changing the degree of uptake of Fgf8 into its target cells, we are able to alter the shape of the Fgf8 gradient. Our results demonstrate that a freely diffusing morphogen can set up concentration gradients in a complex multicellular tissue by a simple source-sink mechanism and that both Fgfr1 and Fgfr4 are likely receptors for Fgf8 signal transduction during zebrafish gastrulation. |
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