RLIP76-dependent suppression of PI3K/AKT/Bcl-2 pathway by miR-101 induces apoptosis in prostate cancer

Highlights

• miR-101 inhibited prostate cancer cell proliferation and enhanced apoptosis.

• miR-101 directly targeted and regulated RLIP76 expression.

• miR-101 suppressed PI3K/Akt/Bcl-2 signaling pathway by targeting RLIP76.

Abstract

MicroRNA-101 (miR-101) participates in carcinogenesis and tumor progression in various cancers. However, its biological functions in prostate cancer are still unclear. Here, we demonstrate that miR-101 represents a critical role in regulating cell apoptosis in prostate cancer cells. We first demonstrated that miR-101 treatment promoted apoptosis in DU145 and PC3 cells by using flow cytometric analysis and transmission electron microscopy (TEM). To verify the mechanisms, we identified a novel miR-101 target, Ral binding protein 1 (RLIP76). We found miR-101 transfection significantly suppresses RLIP76 expression, which can transactivate phosphorylation of PI3K-Akt signaling, and resulted in an amplification of Bcl2-induced apoptosis. Furthermore, we demonstrated that RLIP76 overexpression could reverse the anti-tumor effects of miR-101 in DU145 and PC3 cells by using flow cytometry assay and MTT assay. Taken together, our results revealed that the effect of miR-101 on prostate cancer cell apoptosis was due to RLIP76 regulation of the PI3K/Akt/Bcl-2 signaling pathway.

Introduction

Prostate cancer represents one of the most common cancers in men [1]. With the development of early diagnosis and treatment modalities, the 5-year survival rate of prostate cancer has been improved over the past two decades [2]. However, even with aggressive intervention (including surgery, radiotherapy and chemotherapy), it does not ensure a cure especially for those castration-resistant prostate cancers (CRPC) [3]. To improve the prognosis of patients with malignant prostate cancer, it is urgent to identify new molecular biomarkers that regulate malignant biological behavior of prostate cancer cells.

MicroRNAs (miRNA), a class of nocoding RNA molecules, negatively mediate the translation of mRNAs by interacting with complementary sites in the 3′ untranslated region (3′ UTR) [4]. The close relationship between miRNAs and cellular functions, such as cell growth, migration, and cell cycle, are well documented [5]. They also participate in several pathological processes of cancers [6]. Some miRNAs have been regard as suppressor genes in specific cancers by directly targeting oncogenes [7]. Mounting evidences indicate that microRNA-101 (miR-101) is a potential cancer marker and functions as a tumor suppressor gene in several tumors. Specially, aberrant low expression level of miR-101 has been demonstrated in a variety of malignancies, such as lung, colon and bladder cancers [8], [9], [10]. Furthermore, recent studies have demonstrated that miR-101 decreases the invasiveness and regulates cell proliferation in prostate cancers, highlighting its importance in prostate cancer [11], [12]. However, the molecular mechanisms of miR-101 in cell apoptosis in prostate cancer cells are still unknown.

Ral binding protein 1 (RLIP76), a GTPase-activating protein, is required for tumor progressions, including cell proliferation, apoptosis and invasion [13], [14], [15]. Previous studies have shown that the suppression of PI3K/Akt signaling has been observed together with the downregulation of RLIP76 in pancreatic cancer. Consistent with its growth-promoting role, RLIP76 is regarded as an oncogene [16]. Importantly, excessive expression of RLIP76 is positively correlated with tumor growth in prostate cancer xenografts, suggesting that increased RLIP76 level may be critical to prostate cancer patient survival. Here, we confirmed the function of miR-101 in prostate cancer cell apoptosis and demonstrated that miR-101 suppresses PI3K/Akt pathway via directly targeting the multi-functional transporter protein RLIP76.