YAP1 overexpression is associated with poor prognosis of breast cancer patients and induces breast cancer cell growth by inhibiting PTEN

Abstract

YES-associated protein 1 (YAP1) plays a key role as a transcriptional coactivator in the Hippo tumor suppressor pathway. YAP1 is overexpressed in a variety of cancers and is considered to be encoded by a proto-oncogene. However, the role of YAP1 remains debatable, because both gain and loss of YAP1 expression have both been reported in breast cancer (BC). Here, we found that elevated expression of YAP1 mRNA in BC was negatively correlated with relapse-free, distant metastases-free and overall survival rates. We then knocked down or overexpressed YAP1 in human BC cells, and examined cell proliferation, apoptosis, and tumorigenic ability in vivo. We identified that YAP1 promotes cell growth and inhibits cell apoptosis of BC through the phosphatase and tensin homolog deleted on chromosome 10-AKT signaling pathway, and thus suggest that YAP1 might serve as a new target for inhibiting BC progression.

Keywords: AKT; Hippo; PTEN; YAP1; apoptosis; breast cancer.

Figure 1

YAP1 is upregulated in BC cells and correlated with poor prognosis of BC patients. (A,B) Western blotting of YAP1 expression in normal breast epithelial cells and BC cells. (C–E) Kaplan–Meier plots showing the association between YAP1 mRNA expression and patients’ relapse‐free survival (C), distant metastases‐free survival (D), and overall survival (E). Data are presented as mean ± SD of three biological replicates and are analyzed by two‐tailed Student's t test; *P < 0.05 vs MCF10A. HR, hazard ratio.

Figure 2

YAP1 modulates proliferation of BC cells. (A) Western blot of indicated BC cells transfected with YAP vector, YAP1, YAP1‐RNAi vector, YAP1‐RNAi1 or YAP1‐RNAi2. (B) Protein quantification of the western blot results. (C,D) CCK8 assays revealed that overexpression of YAP1 significantly increased the growth rate of indicated cells. (E,F) CCK8 assays revealed that downregulation of endogenous YAP1 significantly reduced the growth rate. Data are presented as mean ± SD of three biological replicates and are analyzed by two‐tailed Student's t test; *P < 0.05 vs vector, **P < 0.01 vs vector.

Figure 3

(A) Images of excised tumors from five BALB/C nude mice at 42 days after injection with YAP1 RNAi1‐transfected cells and vector transfected cells. (B) Average weight of excised tumors. Data are presented as mean ± SD and are analyzed by two‐tailed Student's t test (n = 5 per group); *P < 0.05 vs vector.

Figure 4

YAP1 affects cell apoptosis and proliferation through regulation of PTEN–AKT signaling. (A) Western blot analysis of PTEN, phosphorylated AKT (p‐AKT), and total AKT protein in the indicated BC cell lines. (B) Flow cytometric assays revealed the role of PTEN‐specific inhibitor bpV(HOpic) in the apoptosis of YAP1‐RNAi1‐transduced cells. (C) CCK8 assays revealed the role of bpV(HOpic) in the proliferation of YAP1‐RNAi1‐transduced cells. (D) Western blot analysis of p‐PTEN, PTEN, p‐AKT, and total AKT protein in bpV (HOpic)‐treated YAP1 silenced cells. Data are presented as mean ± SD of three biological replicates and were analyzed by two‐tailed Student's t test; *P < 0.05 vs vector, **P < 0.01 vs vector.