Islet1-mediated activation of the β-catenin pathway is necessary for hindlimb initiation in mice

Abstract

SUMMARYThe transcriptional basis of vertebrate limb initiation, which is a well-studied system for the initiation of organogenesis, remains elusive. Specifically, involvement of the -catenin pathway in limb initiation, as well as its role in hindlimb-specific transcriptional regulation, are under debate. Here, we show that the -catenin pathway is active in the limb-forming area in mouse embryos. Furthermore, conditional inactivation of -catenin as well as Islet1, a hindlimb-specific factor, in the lateral plate mesoderm results in a failure to induce hindlimb outgrowth. We further show that Islet1 is required for the nuclear accumulation of -catenin and hence for activation of the -catenin pathway, and that the -catenin pathway maintains Islet1 expression. These two factors influence each other and function upstream of active proliferation of hindlimb progenitors in the lateral plate mesoderm and the expression of a common factor, Fgf10. Our data demonstrate that Islet1 and -catenin regulate outgrowth and Fgf10-Fgf8 feedback loop formation during vertebrate hindlimb initiation. Our study identifies Islet1 as a hindlimb-specific transcriptional regulator of initiation, and clarifies the controversy regarding the requirement of -catenin for limb initiation.

ISLET1-Dependent β-Catenin/Hedgehog Signaling Is Required for Outgrowth of the Lower Jaw

“…The nuclear ␤-catenin level in the majority of epithelial cells was diminished in Isl1 Shh-Cre mice (Fig. 6C), consistent with the notion that ISL1 is required for the activation of ␤-catenin signaling (17,20). To test the possibility of ISL1 regulation of the WNT/␤-catenin pathway via repression of the WNT antagonists, we performed chromatin immunoprecipitation (ChIP) assays using mandibular samples at E10.5.…”

“…Previous studies have shown ISL1 activation of ␤-catenin signaling in the development of muscular structures and branchial arch epithelium (17,20). Although these studies have provided the immunohistochemical evidence that ISL1 may regulate ␤-catenin signaling through modulating cellular accumulation of ␤-catenin, we still do not know how ISL1 as a transcription factor regulates the ␤-catenin pathway due to lack of mechanical evidence.…”

“…Previous studies have demonstrated that ISL1 is critical for the nuclear accumulation of ␤-catenin during development (17,20). To assess the role of ISL1 in controlling ␤-catenin signaling in mandibular morphogenesis, we first examined the impact of Isl1 ablation on Wnt/␤-catenin signaling by detecting TOPGAL (Tcf optimal promoter ␤-galactosidase) activity as a Wnt/␤-catenin signaling reporter in the Isl1 Shh-Cre mandible.…”

“…Deletion of ␤-catenin in Isl1-expressing cells resulted in the loss of epithelial Fgf8 expression and death of mesenchymal cells (17). ISL1 is thus postulated to regulate development of the mandible through signaling pathways such as ␤-catenin, FGF, and the BMP regulatory network in mouse development (8,16,17,20). However, the early lethality of homozygous Isl1 mutants at E9.0 prevented the direct analysis of potential roles for Isl1 in the regulation of mandibular signals.…”

Migration of cardiomyocytes is essential for heart regeneration in zebrafish

“…In situ hybridization on sections 14 m thick was performed as previously described (Kawakami et al, 2011). Immunostaining on sections was performed according to a standard procedure Raya et al, 2003).…”