Sustained TGFβ exposure suppresses Smad and non-Smad signalling in mammary epithelial cells, leading to EMT and inhibition of growth arrest and apoptosis

Abstract

To better understand the dual, tumour-suppressive and tumour-promoting function of transforming growth factor-β (TGFβ), we analysed mammary epithelial NMuMG cells in response to short and long-term TGFβ exposure. NMuMG cells became proliferation-arrested and apoptotic after exposure to TGFβ for 2–5 days, whereas surviving cells underwent epithelial–mesenchymal transition (EMT). After chronic TGFβ exposure (2–3 weeks), however, NMuMG cells became resistant to proliferation arrest and apoptosis, showing sustained EMT instead (TD cells). EMT was fully reversed by a pharmacologic TGFβ-receptor-I kinase inhibitor or withdrawal of TGFβ for 6–12 days. Interestingly, both cell cycle arresting/proapoptotic (Smads, p38 kinase) and antiapoptotic, proliferation and EMT-promoting signalling pathways (PI3K–PKB/Akt, ERK) were co-suppressed to low, but significant levels. Except for PI3K-Akt, TGFβ-dependent downregulation of these signalling pathways in transdifferentiated (TD) cells was fully reversed upon TGFβ withdrawal, together with partial re-induction of proliferation arrest and apoptosis. Co-injection of non-tumorigenic NMuMG cells with tumour-forming CHO cells oversecreting exogenous TGFβ1 (CHO-TGFβ1) allowed outgrowth of epithelioid cells in CHO-TGFβ1 cell-induced tumours. These epithelial islands enhanced CHO-TGFβ1 tumour cell proliferation, possibly due to chemokines (for example, JE/MCP-1) secreted by NMuMG/TD cells. We conclude that suppression of antiproliferative, proapoptotic TGFβ signalling in TD cells may permit TGFβ-dependent proliferation, survival and EMT-enhancing signalling pathways to act at low levels. Thus, TGFβ may modulate its own signalling to facilitate switching from tumour suppression to tumour progression.