Planar Cell Polarity Pathways in Breast Development

Epithelial cells are the building blocks of tissues which line surfaces throughout the body. These cells are robustly arranged into patterns and structures depending on the specialised functions of various organs. How single cells arrange into these structures is a fundamental question in biology. The mammary gland provides an ideal system to study epithelial morphogenesis as it develops post-natally and remodels during adult life. Like many internal organs, mammary epithelia are assembled into a branched network of ducts, with milk producing alveoli developing during pregnancy and lactation. However, the mechanisms which regulate the organisation of epithelial cells into distinct morphological structures remain to be uncovered. This study investigated the hypothesis that ductal morphogenesis could be induced by the following 3 ways: a) collective cell migrations along the length of the duct, b) oriented cell divisions and c) regulated cell shape and intercellular adhesions.
Collective cell behaviours are commonly regulated by Planar Cell Polarity (PCP) proteins, the phenomenon by which cells are polarised in the 2D plane of an epithelium.
When cultured in 3D matrix, mouse mammary epithelial cells form the ducts and branches of the mammary gland. Here we investigated alternative 3D mammary culture techniques and designed tools to analyse PCP pathways in primary organoid cultures. These include shRNA knockdown of Vangl2 and insertion of H2B-RFP to analyse oriented cell division in organoids. PCP in the pubertal developing mammary gland was investigated and reveals Vangl2 protein expression mirrors increased ductal elongation and branching, and Vangl2 localisation to some cell membranes, which suggests that Vangl2 plays roles in the development of mammalian mammary epithelia.
Mammary gland morphology was investigated using a 3D immunofluorescence and microscopy technique of whole mount tissue, and we showed a distinct organisation of myoepithelial cells around elongating ducts. By understanding the mechanisms of epithelial morphogenesis under normal conditions, we understand more how these processes could function abnormally in disease states such as cancer.