Ras suppressor-1 promotes apoptosis in breast cancer cells by inhibiting PINCH-1 and activating p53-upregulated-modulator of apoptosis (PUMA); verification from metastatic breast cancer human samples

Abstract

Metastasis, responsible for most deaths from breast cancer (BC), is a multistep process leading to cancer cell spread. Extracellular matrix (ECM)-related adhesion and apoptosis resistance play pivotal role in metastasis. Ras suppressor-1 (RSU-1) localizes to cell-ECM adhesions and binds to pro-survival adhesion protein PINCH-1. Little is known about the role of RSU-1 in BC. In the present study, we investigated the role of RSU-1 in BC metastasis using two BC cell lines that differ in terms of their metastatic potential and a set of 32 human BC samples from patients with or without lymph node metastasis. We show that RSU-1 is upregulated in the aggressive MDA-MB-231 cells compared to MCF-7 and that its silencing by siRNA leads to upregulation of PINCH-1, induction of proliferation and reduction of apoptosis through downregulation of the pro-apoptotic gene p53-upregulated-modulator-of-apoptosis (PUMA). Our findings in the cell lines were further validated in the human BC tissues where normal adjacent tissues were used as controls. We demonstrate for the first time, that RSU-1 expression is upregulated in metastatic BC samples and downregulated in non-metastatic while it is negatively correlated with PINCH-1 and positively correlated with PUMA expression, suggesting that a pro-apoptotic mechanism is in place in metastatic BC samples and identifying RSU-1 as a potentially interesting molecule that needs to be evaluated further as a novel BC metastasis biomarker.

Cancer cell metastasis; perspectives from the focal adhesion

“…In accordance with the cell line data that suggested RSU-1 to be a negative regulator of PINCH-1, PINCH-1 expression showed dramatic reduction in BC samples both at the mRNA and protein level with statistically significant reduction in metastatic samples, in which RSU-1 was found to be elevated. Moreover, the assessment of the pro-apoptotic PUMA expression also confirmed findings in the cell lines showing an increase of PUMA expression in BC and metastatic samples in particular and a positive statistically significant correlation with RSU-1 expression in the same samples [43] . Therefore, RSU-1 should definitely be evaluated further as a potential metastasis biomarker in BC tissue samples.…”

“…Interestingly, the data showed that RSU-1 mRNA expression was significantly increased in BC tissues when compared to normal adjacent tissue. Moreover, RSU-1 expression was significantly reduced in non-metastatic samples and significantly elevated in metastatic samples, clearly stating a different role of RSU-1 depending on the aggressiveness of the tumor, that could potentially lead to the use of RSU-1 as a novel biomarker of metastasis [43] . In accordance with the cell line data that suggested RSU-1 to be a negative regulator of PINCH-1, PINCH-1 expression showed dramatic reduction in BC samples both at the mRNA and protein level with statistically significant reduction in metastatic samples, in which RSU-1 was found to be elevated.…”

“…Moreover, siRNA-mediated gene silencing of RSU-1 expression in both HCC cell lines resulted in increased proliferation, reduced cell adhesion and cell invasion in the aggressive HepG2 cells [42] , suggesting that RSU-1 enhances adhesion and invasion in the aggressive HepG2 cells. Interestingly, in another recent study by our group, RSU-1 was investigated in BC cell lines that differ in terms of their metastatic potential (namely, non-invasive MCF-7 cells and highly invasive MDA-MB-231 cells) as well as in a set of thirty two [32] human BC samples from patients with or without lymph node metastasis [43] . In this study, the findings from the experiments performed in the cell lines were further supported by the findings in the human samples validating the hypothesis that RSU-1 plays a vital role in BC metastasis.…”

Particularly interesting Cys-His-rich protein is highly expressed in human intracranial aneurysms and resists aneurysmal rupture

“…PINCH-1 promotes B-cell lymphoma (Bcl)-2-dependent survival signalling and inhibits c-Jun N-terminal kinase (JNK)-mediated apoptosis in the PrE (13). Although the function of PINCH remains unknown, increasing numbers and accumulating evidence indicates that PINCH has been correlated with cancer development, invasion and metastasis in malignant cells, including breast cancer (14), pseudomyxoma peritonei (15), gastric adenocarcinoma (16) and rectal cancer (17). For example, PINCH is demonstrated to protect tumor cells from apoptosis by increasing the activity of the pro-survival protein extracellular signal-regulated kinase (ERK)1/2 and Akt (18).…”

Signaling via PINCH: Functions, binding partners and implications in human diseases

“…This result contradicts, at least in part, with the finding that Rsu1 stabilized PINCH1 29 , further complicating the functional relationship between PINCH1 and Rsu1.…”