Polarity as a Regulator of Metaplasia

Abstract

Cell polarity is an important regulator of cellular processes and is vital in helping to prevent metaplasia and tumorigenesis. There are three many polarity complexes that regulate and maintain epithelial cellular polarity. The Par and Crumbs complexes locate to the apical membrane of the cell, while the Scribble complex is located basolaterally. Of the Scribble complex components, the polarity protein Hugl1, also known as Mgl1 in mice, is especially important in helping to maintain apical basolateral and planar polarity, and is lost in multiple types of cancer. When Hugl1 expression is lost in epithelial cells, it results in a mesenchymal phenotype. We now show that the loss of Hugl1 fundamentally shifts the cellular phenotype and specifically alters EGFR trafficking and signaling. Loss of Hugl1 results in the nuclear translocation of Taz and Slug, increased migration, and the mislocalization of EGFR (Epidermal Growth Factor Receptor), driving cellular growth. Hugl1 regulates the expression of multiple cell identity markers and its loss results in stem cell characteristics, including the increased expression of CD44, and a decrease of CD49f and CD24 expression. The loss of Hugl1 also results in increased growth in soft-agar and prolonged survival when transplanted into NOD-SCID mice; its loss also results in EGF-dependent migration which aids in increasing mammosphere survival. Furthermore, isolated EGFR mislocalization via a point mutation (P667A) also drives these same phenotypes, including activation of Akt and Taz nuclear translocation, indicating the importance of Hugl1 in the regulation of EGFR localization and its signaling. In mice, the loss of total Mgl1 is lethal within days of birth due to hydrocephaly and results in the formation of rosette like structures in the brain that are reminiscent of neuroectodermal tumors. We designed a targeted Mgl1 knockout in the mammary epithelial cells using the Cre/Lox system to evaluate the effects of Mgl1 loss in murine mammary gland development and tumorigenesis. The loss of Mgl1 expression in mice inhibits ductal outgrowth, increases side branching and epithelial layers, and results in the mislocalization of EGFR. While overt mammary tumors did not develop, some individuals did develop hyperplastic nodules that could progress into cancer. The knockdown of Hugl1 in vitro and Mgl1 in vivo reveal how the loss of polarity and presence of Hugl1 results in cancer stem cell characteristics, increased migration, and abnormal signaling due to the mislocalization of EGFR. While these changes result in metaplasia and a potential pre-cancerous state, the loss of Hugl1 alone is not enough to drive the cancer progression, indicating that other mutations or factors are necessary for the development of breast cancer. Because of the key role polarity plays in the prevention of breast cancer development we investigated if the addition of Hugl1 back into breast cancer cells could revert the cancerous cells to a normal epithelial phenotype. Most of the breast cancer cells transfected with Hugl1 expression did not survive, indicating that the re-expression of polarity regulators forces cancer cells to die. The small percentage of cells that did survive re-expression of Hugl1 had retarded growth in soft agar and a decrease in EGFR expression. Together, these data indicate that Hugl1 expression and EGFR activity are closely related and that Hugl1 is required for the proper localization and signaling of EGFR. When Hugl1 is lost, EGFR is mislocalized and fails to be degraded properly, promoting pre-neoplastic changes.