The canonical Wnt/ß-catenin signaling pathway regulates Fgf signaling for early facial development

Abstract

The canonical Wnt/β-catenin signaling pathway has implications in early facial development; yet, its function and signaling mechanism remain poorly understood. We report here that the frontonasal and upper jaw primordia cannot be formed after conditional ablation of β-catenin with Foxg1-Cre mice in the facial ectoderm and the adjacent telencephalic neuroepithelium. Gene expression of several cell-survival and patterning factors, including Fgf8, Fgf3, and Fgf17, is dramatically diminished in the anterior neural ridge (ANR, a rostral signaling center) and/or the adjacent frontonasal ectoderm of the β-catenin conditional mutant mice. In addition, Shh expression is diminished in the ventral telencephalon of the mutants, while Tcfap2a expression is less affected in the facial primordia. Apoptosis occurs robustly in the rostral head tissues following inactivation of Fgf signaling in the conditional mutants. Consequently, the upper jaw, nasal, ocular and telencephalic structures are absent, but the tongue and mandible are relatively developed in the conditional mutants at birth. Using molecular biological approaches, we demonstrate that the Fgf8 gene is transcriptionally targeted by Wnt/β-catenin signaling during early facial and forebrain development. Furthermore, we show that conditional gain-of-function of β-catenin signaling causes drastic upregulation of Fgf8 mRNA in the ANR and the entire facial ectoderm, which also arrests facial and forebrain development. Taken together, our results suggest that canonical Wnt/β-catenin signaling is required for early development of the mammalian face and related head structures, which mainly or partly acts through the initiation and modulation of balanced Fgf signaling activity.

LDL Receptor–Related Protein 6 Modulates Ret Proto-Oncogene Signaling in Renal Development and Cystic Dysplasia

“…Wholemount in situ hybridization was carried out as previously described, 48 using the DIG-labeled cRNA riboprobes (Supplemental Table 1). Samples were photographed using a Carl Zeiss stereological microscope (Lumar V12).…”

β-catenin regulates Pax3 and Cdx2 for caudal neural tube closure and elongation

“…Among the crucial tail bud patterning genes, T, Tbx6 and Fgf8 have been identified as the downstream targets of Wnt/β-catenin signaling in the tail bud or in the anterior neural ridge (Szeto and Kimelman, 2004;Wang et al, 2011;Yamaguchi et al, 1999b). Because these genes are not altered at E9.5 in the tail buds of the mutants, we then examined these genes at a later stage when neural tube closure is completed in wild-type or doubleheterozygous embryos.…”

“…X-gal staining and wholemount in situ hybridization Embryos were fixed in 1% paraformaldehyde (PFA) for ~30 minutes on ice and processed for X-gal staining as described previously (Song et al, 2009;Wang et al, 2011). Embryos fixed in 4% PFA overnight at 4°C were processed for wholemount in situ hybridization using digoxigenin-labeled antisense RNA probes (supplementary material Cre/+ ) embryos were used for the normal controls, which showed no significant differences in X-gal staining or in situ mRNA signals.…”

“…4F). Similar to previously described studies (Song et al, 2009;Wang et al, 2011), L cells were transiently transfected with the wild-type or mutated promoter-driven luciferase reporters, together with the constitutively active β-catenin and wild-type Lef1 expression constructs. A control expression vector (pcDNA) was introduced into L cells to monitor the baseline luciferase reporter activity.…”

“…Chromatin extraction and immunoprecipitation were performed as previously described (Song et al, 2009;Wang et al, 2011). Anti-β-catenin antibody (Santa Cruz Biotech; sc-7199) was used to pull down the β-catenin-Tcf/Lef DNA-binding complex, and rabbit IgG (Invitrogen; 10500C) was used for the negative control.…”