ZEB1 Imposes a Temporary Stage-Dependent Inhibition of Muscle Gene Expression and Differentiation via CtBP-Mediated Transcriptional Repression

Abstract

Skeletal muscle development is orchestrated by the myogenic regulatory factor MyoD, whose activity is blocked in myoblasts by proteins preventing its nuclear translocation and/or binding to G/C-centered E-boxes in target genes. Recent evidence indicates that muscle gene expression is also regulated at the cis level by differential affinity for DNA between MyoD and other E-box binding proteins during myogenesis. MyoD binds to G/C-centered E-boxes, enriched in muscle differentiation genes, in myotubes but not in myoblasts. Here, we used cell-based and in vivo Drosophila, Xenopus laevis, and mouse models to show that ZEB1, a G/C-centered E-box binding transcriptional repressor, imposes a temporary stage-dependent inhibition of muscle gene expression and differentiation via CtBP-mediated transcriptional repression. We found that, contrary to MyoD, ZEB1 binds to G/C-centered E-boxes in muscle differentiation genes at the myoblast stage but not in myotubes. Its knockdown results in precocious expression of muscle differentiation genes and acceleration of myotube formation. Inhibition of muscle genes by ZEB1 occurs via transcriptional repression and involves recruitment of the CtBP corepressor. Lastly, we show that the pattern of gene expression associated with muscle differentiation is accelerated in ZEB1^−/− mouse embryos. These results set ZEB1 as an important regulator of the temporal pattern of gene expression controlling muscle differentiation.

ACKNOWLEDGMENTS

We are indebted to researchers who generously provided us with reagents (see Materials and Methods) and especially to Y. Higashi (Institute for Development Research, Kasugai, Japan) for the δEF1^+/− (ZEB1^+/−) mice. We are also grateful to J. Skeath (Washington University School of Medicine, St. Louis, MO) for help in Drosophila experiments. We apologize to researchers whose work was cited indirectly through reviews due to space limitations.

The different parts of this study were independently funded by grants from the Spanish Ministry of Economy and Competitiveness (formerly MICINN) (grants BFU2007-60302 and BFU2010-15163), the La Caixa Foundation (LCF), the AVON-SAU Breast Cancer Research Campaign, the Olga Torres Foundation (FOT), the Spanish Association against Cancer (AECC), and the European Commission to A.P. L.S. is the recipient of a Ph.D. scholarship from the Ministry of Education, Culture and Sports (FPU Program, scholarship AP2010-4495), and her salary was previously partly funded by the AECC and FOT. E.S.-T.'s salary was partly funded by the LCF and CIBERehd. We declare that we have no competing financial interests.