RESUMO
The Wnt pathway activates target genes by controlling the ß-catenin-T-cell factor (TCF) transcriptional complex during embryonic development and cancer. This pathway can be potentiated by R-spondins, a family of proteins that bind RNF43/ZNRF3 E3 ubiquitin ligases and LGR4/5 receptors to prevent Frizzled degradation. Here we demonstrate that, during Xenopus anteroposterior axis specification, Rspo2 functions as a Wnt antagonist, both morphologically and at the level of gene targets and pathway mediators. Unexpectedly, the binding to RNF43/ZNRF3 and LGR4/5 was not required for the Wnt inhibitory activity. Moreover, Rspo2 did not influence Dishevelled phosphorylation in response to Wnt ligands, suggesting that Frizzled activity is not affected. Further analysis indicated that the Wnt antagonism is due to the inhibitory effect of Rspo2 on TCF3/TCF7L1 phosphorylation that normally leads to target gene activation. Consistent with this mechanism, Rspo2 anteriorizing activity has been rescued in TCF3-depleted embryos. These observations suggest that Rspo2 is a context-specific regulator of TCF3 phosphorylation and Wnt signaling.
Assuntos
Padronização Corporal/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Fator 3 de Transcrição/antagonistas & inibidores , Via de Sinalização Wnt/efeitos dos fármacos , Proteínas de Xenopus/antagonistas & inibidores , Proteínas de Xenopus/fisiologia , Animais , Padronização Corporal/fisiologia , Embrião não Mamífero/anormalidades , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Genes Reporter , Cabeça/embriologia , Fosforilação/efeitos dos fármacos , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Fator 3 de Transcrição/metabolismo , Proteínas de Xenopus/biossíntese , Proteínas de Xenopus/genética , Proteínas de Xenopus/metabolismo , Proteínas de Xenopus/farmacologia , Xenopus laevis/embriologiaRESUMO
Several E-box-binding transcription factors regulate individual and collective cell migration and enhance the motility of epithelial cells by promoting epithelial-mesenchymal transition (EMT). Here, we characterized the role of a subset of these transcription factors and the EMT proteome in branching morphogenesis of mammary epithelial tissues using a three-dimensional organotypic culture model of the mammary duct. We found that the transcription factors Snail1, Snail2, and E47 were transiently upregulated at branch sites; decreasing the expression of these transcription factors inhibited branching. Conversely, ectopic expression of Snail1, Snail2, and E47 induced branching in the absence of exogenous stimuli. These changes correlated with the expression of mesenchymal markers and repression of E-cadherin, which was essential for branching. Snail1 and Snail2 also promoted cell survival at branch sites, but this was not sufficient to induce branching. These findings indicate that Snail1, Snail2, and E47 can promote collective migration during branching morphogenesis of mammary epithelial tissues through key regulators of EMT.
