Yap1 promotes the survival and self-renewal of breast tumor initiating cells via inhibiting Smad3 signaling

Tumor initiating cells (TICs) serve as the root of tumor growth. After identifying TICs in spontaneous breast tumors of the MMTV-Wnt1 mouse model, we confirmed the specific expression and activation of Yes-associated protein 1 (Yap1) within TICs. To investigate the role of Yap1 in the self-renewal of breast TICs and the underlying mechanism, we sorted CD49f(high)EpCAM(low) cells as breast TICs. Active Yap1 with ectopic expression in breast TICs promoted their colony formation in vitro (p<0.01) and self-renewal in vivo (p<0.01), and led to a 4-fold increase in TIC frequency (p<0.05). A conditional knock-out mouse was reconstructed to generate Yap1 knock-out breast tumors. The loss of Yap1 led to a dramatic growth disadvantage of breast TICs in vitro (p<0.01) and in vivo (p<0.01), and it also led to an over 200-fold decrease in TIC frequency (p<0.01). The expression of active Yap1 was negatively correlated with that of phosphorylated Smad3 (p-Smad3). Transforming growth factor beta (TGF-beta) served as a strong enhancer of Smad3 and an inhibitor of clonogenesis of TICs. The presence of SIS3, a specific inhibitor of Smad3, could rescue the TGF-beta-induced growth inhibition and reverse the Smad3 inhibition by Yap1. Analysis of a database containing 2,072 human breast cancer samples showed that higher expressions of Yap1 correlated with a poorer outcome of a 15-year survival rate and median overall survival (mOS) in patients, especially in those with basal breast tumors without estrogen receptor 1 (ER) expression. The findings indicate that active Yap1 promotes the self-renewal of breast TICs by inhibiting Smad3 signaling.