Loss of epigenetic regulation disrupts lineage integrity, reactivates multipotency and promotes breast cancer

Systematically investigating the scores of genes mutated in cancer and discerning real drivers from inconsequential bystanders is a prerequisite for Precision Medicine, but remains challenging. Here, we developed a somatic CRISPR/Cas9 mutagenesis screen to study 215 recurrent long-tail breast cancer genes, which revealed epigenetic regulation as major tumor suppressive mechanism. We report that core or accessory components of the COMPASS histone methylase complex including KMT2C, KDM6A, BAP1 and ASXL2 (EpiDrivers) cooperate with PIK3CAH1047R to transform mouse and human breast epithelial cells. Mechanistically, we find that Cre-mediated activation of PIK3CAH1047R elicited an aberrant alveolar lactation program in luminal cells, which was exacerbated upon loss of EpiDrivers. Remarkably, EpiDriver loss in basal cells also triggered an alveolar-like lineage conversion and accelerated formation of luminal-like tumors, suggesting a basal origin for luminal tumors. As EpiDrivers are mutated in 39% of human breast cancers, lineage infidelity and lactation mimicry may significantly contribute to early steps of breast cancer progression.

Extracellular matrix stiffness regulates degradation of the Hippo kinase MST2 via SCF βTrCP

Tumor microenvironments display disrupted mechanical properties, including altered extracellular matrix (ECM) rigidity. ECM stiffening perturbs cell tensional homeostasis resulting in activation of mechanosensing transcriptional co-activators, such as the Hippo pathway effectors YAP and TAZ. The Hippo pathway plays central roles in development and tumorigenesis, but how the proteostasis of the Hippo kinase MST2 is regulated remains unknown. We show that MST2 levels decrease upon changes in ECM rigidity via proteasome degradation. MST2 degradation is enhanced in human breast epithelial cells that are cultured in stiffer microenvironments due to integrin and integrin-linked kinase activation. MST2 knockdown resulted in increased nucleus-to-cytoplasm ratio of YAP, increased proliferation rates in a soft microenvironment and F-actin alignment in cells cultured in intermediate stiffness. We found that MST2 is ubiquitinated by the SCFβTrCP ubiquitin ligase, and site-directed mutagenesis combined with computational molecular dynamics studies revealed that βTrCP binds MST2 via a noncanonical degradation motif. Our study uncovers the underlying biochemical mechanisms controlling MST2 degradation and demonstrates how alterations in the microenvironment rigidity regulate the proteostasis of a central Hippo pathway component.