Mammary branch initiation and extension are inhibited by separate pathways downstream of TGFβ in culture

During the branching morphogenesis process that builds epithelial trees, signaling from stimulatory and inhibitory growth factors is integrated to control branch initiation and extension into the surrounding stroma. Here, we examined the relative roles played by these stimulatory and inhibitory signals in the patterning of branch initiation and extension of model mammary epithelial tubules in culture. We found that although several growth factors could stimulate branching, they did not determine the sites at which new branches formed or the lengths to which branches extended. Instead, branch initiation and extension were defined by two separate signals downstream of the inhibitory morphogen, transforming growth factor (TGF)-β. Branch initiation was controlled by signaling through p38 mitogen-activated protein kinase, whereas branch extension was controlled by Smad-mediated induction of a second diffusible inhibitor, Wnt5a. These data suggest that mammary epithelial branching is patterned predominately by repulsive signaling, and that TGFβ activates multiple inhibitory pathways to refine the architecture of the tree.