Notch1 transactivates Notch3 to drive terminal differentiation in stratified squamous epithelia. Notch1 and other Notch receptor paralogs cooperate to act as a tumor suppressor in squamous cell carcinomas (SCCs). However, Notch1 can be stochastically activated to promote carcinogenesis in murine models of SCC. Activated form of Notch1 promotes xenograft tumor growth when expressed ectopically. Here, we demonstrate that Notch1 activation and epithelial–mesenchymal transition (EMT) are coupled to promote SCC tumor initiation in concert with transforming growth factor (TGF)-β present in the tumor microenvironment. We find that TGFβ activates the transcription factor ZEB1 to repress Notch3, thereby limiting terminal differentiation. Concurrently, TGFβ drives Notch1-mediated EMT to generate tumor initiating cells characterized by high CD44 expression. Moreover, Notch1 is activated in a small subset of SCC cells at the invasive tumor front and predicts for poor prognosis of esophageal SCC, shedding light upon the tumor promoting oncogenic aspect of Notch1 in SCC.

Accumulation of tumor-associated macrophages (TAMs) associates with malignant progression in cancer. However, the mechanisms that drive the pro-tumor functions of TAMs are not fully understood. ZEB1 is best known for driving an epithelial-to-mesenchymal transition (EMT) in cancer cells to promote tumor progression. However, a role for ZEB1 in macrophages and TAMs has not been studied. Here we describe that TAMs require ZEB1 for their tumor-promoting and chemotherapy resistance functions in a mouse model of ovarian cancer. Only TAMs that expressed full levels of accelerated tumor growth. Mechanistically, ZEB1 expression in TAMs induced their polarization toward an F4/80 pro-tumor phenotype, including direct activation of In turn, expression of ZEB1 by TAMs induced and a mesenchymal/stem-like phenotype in cancer cells. In human ovarian carcinomas, TAM infiltration and CCR2 expression correlated with ZEB1 in tumor cells, where along with CCL2 and CD74 determined poorer prognosis. Importantly, ZEB1 in TAMs was a factor of poorer survival in human ovarian carcinomas. These data establish ZEB1 as a key factor in the tumor microenvironment and for maintaining TAMs' tumor-promoting functions.

Epithelial-to-mesenchymal transition (EMT)-inducing transcription factors (TF) are well known for their ability to induce mesenchymal states associated with increased migratory and invasive properties. Unexpectedly, nuclear expression of the EMT-TF ZEB2 in human primary melanoma has been shown to correlate with reduced invasion. We report here that ZEB2 is required for outgrowth for primary melanomas and metastases at secondary sites. Ablation of Zeb2 hampered outgrowth of primary melanomas in vivo, whereas ectopic expression enhanced proliferation and growth at both primary and secondary sites. Gain of Zeb2 expression in pulmonary-residing melanoma cells promoted the development of macroscopic lesions. In vivo fate mapping made clear that melanoma cells undergo a conversion in state where ZEB2 expression is replaced by ZEB1 expression associated with gain of an invasive phenotype. These findings suggest that reversible switching of the ZEB2/ZEB1 ratio enhances melanoma metastatic dissemination.Significance: ZEB2 function exerts opposing behaviors in melanoma by promoting proliferation and expansion and conversely inhibiting invasiveness, which could be of future clinical relevance.

The mechanisms linking muscle injury and regeneration are not fully understood. Here we report an unexpected role for ZEB1 regulating inflammatory and repair responses in dystrophic and acutely injured muscles. ZEB1 is upregulated in the undamaged and regenerating myofibers of injured muscles. Compared to wild-type counterparts, Zeb1-deficient injured muscles exhibit enhanced damage that corresponds with a retarded p38-MAPK-dependent transition of their macrophages towards an anti-inflammatory phenotype. Zeb1-deficient injured muscles also display a delayed and poorer regeneration that is accounted by the retarded anti-inflammatory macrophage transition and their intrinsically deficient muscle satellite cells (MuSCs). Macrophages in Zeb1-deficient injured muscles show lower phosphorylation of p38 and its forced activation reverts the enhanced muscle damage and poorer regeneration. MuSCs require ZEB1 to maintain their quiescence, prevent their premature activation following injury, and drive efficient regeneration in dystrophic muscles. These data indicate that ZEB1 protects muscle from damage and is required for its regeneration.

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