Hyperactive WNT/CTNNB1 signaling induces a competing cell proliferation and epidermal differentiation response in the mouse mammary epithelium

Abstract

In the past forty years, the WNT/CTNNB1 signaling pathway has emerged as a key player in mammary gland development and homeostasis. While also evidently involved in breast cancer, much unclarity continues to surround its precise role in mammary tumor formation and progression. This is largely due to the fact that the specific and direct effects of hyperactive WNT/CTNNB1 signaling on the mammary epithelium remain unknown. Here we use a primary mouse mammary organoid culture system to close this fundamental knowledge gap. We show that hyperactive WNT/CTNNB1 signaling induces competing cell proliferation and differentiation responses. While proliferation is dominant at lower levels of WNT/CTNNB1 signaling activity, higher levels cause reprogramming towards an epidermal cell fate. We show that this involves de novo activation of the epidermal differentiation cluster (EDC) locus and we identify master regulatory transcription factors that likely control the process. This is the first time that the molecular and cellular dose-response effects of WNT/CTNNB1 signaling in the mammary epithelium have been dissected in such detail. Our analyses reveal that the mammary epithelium is exquisitely sensitive to small changes in WNT/CTNNB1 signaling and offer a mechanistic explanation for the squamous differentiation that is observed in some WNT/CTNNB1 driven tumors.

In vivo imaging of mammary epithelial cell dynamics in response to lineage-biased Wnt/β-catenin activation

“…5F ). In support, analogous phenotypes were recently observed in mammary organoid cultures treated with high-dose CHIR99021, a potent activator of the Wnt/β-catenin pathway ( Mourao et al, 2021 ). Based on our findings, we believe that constitutive Wnt activity in LC initially drives their conversion to basal-like cells, by repressing K8 expression and inducing basal traits such as K5/14, Id4 and p63 expression, and that they subsequently enter the same program of squamous transdifferentiation as mutant BC.…”

“…Indeed, our data showed that these early, b-catenin-induced changes to epithelial cell organization and behavior reflect the progressive transdifferentiation of mammary cells to an HF/epidermal-like fate, marked by the acquisition of HF and IFE marker expression (Figure 5F). In support, analogous phenotypes were recently observed in mammary organoid cultures treated with high-dose CHIR99021, a potent activator of the Wnt/b-catenin pathway (Mourao et al, 2021). Based on our findings, we believe that constitutive Wnt activity in LCs initially drives their conversion to basal-like cells, by repressing K8 expression and inducing basal traits, such as K5/14, Id4, and p63 expression, and that they subsequently enter the same program of squamous transdifferentiation as mutant BCs.…”

Wnt target gene Ascl4 is dispensable for skin appendage development

“…Different levels of Wnt/β-catenin signaling often result in the activation of distinct target genes which in turn determine different cell fates in a context-dependent manner (Matos et al, 2020;Mourao et al, 2021;Söderholm and Cantù, 2021). Our data suggest that Ascl4 expression may require high levels of Wnt/β-catenin activity.…”

Wnt target gene Ascl4 is dispensable for skin appendage development

“…Different levels of Wnt/β-catenin signaling often result in the activation of distinct target genes which in turn determine different cell fates in a context-dependent manner (Matos et al, 2020; Mourao et al, 2021; Söderholm & Cantù, 2021). Our data suggest that Ascl4 expression may require high levels of Wnt/β-catenin activity.…”