A bi-modal function of Wnt signalling directs an FGF activity gradient to spatially regulate neuronal differentiation in the midbrain

Abstract

FGFs and Wnts are important morphogens during midbrain development, but their importance and potential interactions during neurogenesis are poorly understood. We have employed a combination of genetic and pharmacological manipulations in zebrafish to show that during neurogenesis FGF activity occurs as a gradient along the anterior-posterior axis of the dorsal midbrain and directs spatially dynamic expression of the Hairy gene her5. As FGF activity diminishes during development, Her5 is lost and differentiation of neuronal progenitors occurs in an anterior-posterior manner. We generated mathematical models to explain how Wnt and FGFs direct the spatial differentiation of neurons in the midbrain through Wnt regulation of FGF signalling. These models suggested that a negative-feedback loop controlled by Wnt is crucial for regulating FGF activity. We tested Sprouty genes as mediators of this regulatory loop using conditional mouse knockouts and pharmacological manipulations in zebrafish. These reveal that Sprouty genes direct the positioning of early midbrain neurons and are Wnt responsive in the midbrain. We propose a model in which Wnt regulates FGF activity at the isthmus by driving both FGF and Sprouty gene expression. This controls a dynamic, posteriorly retracting expression of her5 that directs neuronal differentiation in a precise spatiotemporal manner in the midbrain.

Stathmin-like 4 is critical for the maintenance of neural progenitor cells in dorsal midbrain of zebrafish larvae

“…Canonical Wnt signaling is one of the key signal to mediate midbrain development by regulating proliferation and morphogenesis with a bi-modal control upon FGF activity78. To understand whether Wnt morphogens may regulate stmn4 , we suppressed Wnt activity by applying transgenic fish Tg(hsp70l:dkk1-GFP) w32 to overexpress dkk1 , a Wnt signaling inhibitor, and to examine the expression of lef1 (a Wnt-activated downstream gene) and stmn4 (Fig.…”

“…Mutual interactions between Wnts and fibroblast growth factors (Fgfs) originated from the MHB are proposed to control patterning, cell fate decision and proliferation567. The Fgf gradient thus induces the expression of a hairy gene her5 which prevents neuron differentiation invading from anterior brain via cell cycle regulation8 and inhibition of proneural gene expressions9. Although we know some of the core molecular switches mediating cell cycle exit during neuronal differentiation10 yet how the altered mitosis could direct cell fate determination is still far from clear.…”

Temporally coordinated signals progressively pattern the anteroposterior and dorsoventral body axes

“…During CNS development, FGF maintains the midbrain-hindbrain boundary [48,49] and induces caudal cell fates like the hindbrain and spinal cord [50–55]. Unlike Wnt signaling, FGF is not sufficient to ectopically induce caudal cell fates in the forebrain [35] or in animal explants [30].…”

“…Although this supports a more prominent role for Wnt signaling in specifying the broad subdivisions of the CNS [12], it is notable that FGF is required to generate a permissive environment for the caudalizing activity of Wnt [30]. The complex relationship between FGF and Wnt during neural patterning remains to be fully characterized, but a recent study suggests that Wnt may regulate Sprouty expression, providing a mechanism to coordinate Wnt and FGF signaling [55]. …”

“…Evidence indicates that FGF functions as a morphogen, differentially activating posterior genes in a concentration-dependent manner [52,55,56]. Studies of tagged FGF and single molecule fluorescence correlation spectroscopy (FCS) suggest that the FGF gradient is generated by free diffusion of the ligand and receptor-mediated endocytosis [56,57].…”