The mTOR signaling pathway regulates key steps of mammary gland organoid genesis in a temporal manner.

The development of the mammary gland is a complex process that evolves throughout life. It can undergo drastic changes to support lactation, before involuting back to a rudimentary organ after weaning, in a perfectly orchestrated mechanism. This study aimed to identify the pathways coordinating mammary gland organogenesis, using mouse organoids as a model. In developmental assays, the mechanistic target of rapamycin (mTOR) was shown to be a regulator of cellular lineage determination and branching morphogenesis, acting in a time-dependent manner to control these processes. Indeed, mTOR inhibition during the initial growth phase of organoids abrogated the presence of basal epithelial cells, forcing the expansion of the luminal compartment. At later time points during development, mTOR inhibition promoted branching morphogenesis, increasing the organoid capacity to generate branching/buds. Mechanistically, the mTOR signalling inhibition led to alterations in the expression levels of genes and proteins connected to branching morphogenesis, extracellular matrix remodelling, metabolism, and cell migration. Altogether, this study demonstrates the regulatory functions of mTOR in controlling mammary epithelial cells’ capacity to generate organoids.