FGF8 acts as a classic diffusible morphogen to pattern the neocortex

Abstract

SUMMARYGain-and loss-of-function experiments have demonstrated that a source of fibroblast growth factor (FGF) 8 regulates anterior to posterior (A/P) patterning in the neocortical area map. Whether FGF8 controls patterning as a classic diffusible morphogen has not been directly tested. We report evidence that FGF8 diffuses through the mouse neocortical primordium from a discrete source in the anterior telencephalon, forms a protein gradient across the entire A/P extent of the primordium, and acts directly at a distance from its source to determine area identity. FGF8 immunofluorescence revealed FGF8 protein distributed in an A/P gradient. Fate-mapping experiments showed that outside the most anterior telencephalon, neocortical progenitor cells did not express Fgf8, nor were they derived from Fgf8-expressing cells, suggesting that graded distribution of FGF8 results from protein diffusion from the anterior source. Supporting this conclusion, a dominant-negative high-affinity FGF8 receptor captured endogenous FGF8 at a distance from the FGF8 source. New FGF8 sources introduced by electroporation showed haloes of FGF8 immunofluorescence indicative of FGF8 diffusion, and surrounding cells reacted to a new source of FGF8 by upregulating different FGF8-responsive genes in concentric domains around the source. Reducing endogenous FGF8 with the dominant-negative receptor in the central neocortical primordium induced cells to adopt a more posterior area identity, demonstrating long-range area patterning by FGF8. These observations support FGF8 as a classic diffusible morphogen in neocortex, thereby guiding future studies of neocortical pattern formation.

The cortical hem regulates the size and patterning of neocortex

“…FGF8 positively regulates all three PEA3 genes, Etv1, Etv4 and Etv5, which show a nested expression along the medial telencephalon that follows the R/C FGF8 gradient (Fukuchi-Shimogori and Grove, 2003;Toyoda et al, 2010). Electroporation of Wnt3a decreased expression of all three genes (Fig.…”

“…The RTO directs formation of the neocortical area map (Assimacopoulos et al, 2012;Garel et al, 2003;Toyoda et al, 2010), but no equivalently broad role has been established for the hem (Galceran et al, 2000;Yoshida et al, 2006). Yet the hem resembles in position and constituent signaling molecules a powerful patterning source in the caudal neural tube, the roofplate.…”

Essential basal cytonemes take up Hedgehog in the Drosophila wing imaginal disc

“…cDNAs into living embryos (Sanders et al, 2013;Toyoda et al, 2010), by injection of in vitro-synthesized mRNAs into embryos (Muller et al, 2012), and from transgenes under heterologous regulation such as the Gal4-UAS system (Callejo et al, 2011;Kicheva et al, 2007;Zhou et al, 2012). Protein dispersing from expressing cells has been detected by fluorescent imaging, but because expression at an ectopic site can have unforeseen consequences if the systems that normally process and release the morphogen are overwhelmed, or if subcellular structures that are involved are influenced by expression levels, even these direct measures can be problematic in contexts of overexpression and abnormal regulation.…”

Kinetics of receptor occupancy during morphogen gradient formation

“…2 An important part of necessary coordination is provided by morphogens -form-generating molecules that act as regulators of intracellular processes. 3 Morphogens are typically extracellular proteins that are secreted from localized positions within the epithelium and control cell behaviors through binding and activation of cell surface receptors. 4,5 While the mechanisms by which morphogens control cell dynamics are far from being completely understood, for the purposes of this paper it suffices to say that cells exposed to different concentrations of a morphogen have different numbers of occupied cell surface receptors and behave differently.…”

“…Understanding of the effects of morphogens requires analysis of the dynamics of ligand-receptor complexes. [6][7][8] Several studies suggest that morphogens move through tissues in a manner consistent with Fickian diffusion, with effective diffusivity D. 3,[9][10][11][12][13] Their binding to surface receptors is reversible and is commonly modeled by two first order reactions with the rate constants k on and k off . A morphogen molecule bound to a receptor can either dissociate, and continue its diffusion through the tissue, or be internalized into the cell through a process called endocytosis, which effectively terminates the spread of the morphogen from the source of its production.…”