TGFβ selects for pro-stemness over pro-invasive phenotypes during cancer cell epithelial-mesenchymal transition

Transforming growth factor β (TGFβ) induces epithelial-mesenchymal transition (EMT), which correlates with stemness and invasiveness. Mesenchymal-epithelial transition (MET) is induced by TGFβ withdrawal and correlates with metastatic colonization. Whether TGFβ promotes stemness and invasiveness simultaneously via EMT remains unclear. We established a breast cancer cell model expressing red fluorescent protein (RFP) under the E-cadherin promoter. In 2D cultures, TGFβ induced EMT, generating RFP^low cells with a mesenchymal transcriptome, and regained RFP, with an epithelial transcriptome, after MET induced by TGFβ withdrawal. RFP^low cells generated robust mammospheres, with epithelio-mesenchymal cell surface features. Mammospheres that were forced to adhere generated migratory cells, devoid of RFP, a phenotype which was inhibited by a TGFβ receptor kinase inhibitor. Further stimulation of RFP^low mammospheres with TGFβ suppressed the generation of motile cells, but enhanced mammosphere growth. Accordingly, mammary fat-pad-transplanted mammospheres, in the absence of exogenous TGFβ treatment, established lung metastases with evident MET (RFP^high cells). In contrast, TGFβ-treated mammospheres revealed high tumour-initiating capacity, but limited metastatic potential. Thus, the biological context of partial EMT and MET allows TGFβ to differentiate between pro-stemness and pro-invasive phenotypes.