Knockdown of regulator of cullins-1 (ROC1) expression induces bladder cancer cell cycle arrest at the G2 phase and senescence.

Wei Wang,Ping Qu,Yifeng Guo,Jianxin Qiu,Zhengdong Zhou,Jie Fan,Yigang Zeng,Zhihong Liu,Yong Liu

doi:10.1371/journal.pone.0062734

Abstract

Regulator of Cullins-1 (ROC1) is a key subunit in the Cullin-RING ligase (CRL) protein complex. Overexpression of ROC1 protein is associated with tumor progression and poor prognosis of non-muscle invasive bladder transitional cell carcinoma (NMIBC). This study was designed to assess the effects of ROC1 knockdown in bladder cancer cells and to determine the potential mechanisms involved. A total of 112 bladder cancer tissue specimens were recruited for immunohistochemical analyses of ROC1 overexpression. Bladder cancer cell lines were used to knockdown ROC1 expression using ROC1 siRNA. Our data showed that ROC1 knockdown remarkably inhibited bladder cancer cell growth, arrested cells at the G2 phase of the cell cycle, and induced the p53-dependent cell senescence. Molecularly, G2 arrest was associated with upregulation of p21, p27, cyclin B1, and Cdc2 proteins. ROC1 knockdown induced-senescence functioned through p53/p21 pathway. Knockdown of p21 expression partially rescued ROC1 knockdown-induced growth inhibition in cancer cells. Furthermore, nude mouse xenograft analyses confirmed these in vitro data. In conclusion, data from the current study indicate that ROC1 plays an essential role in bladder cancer progression and could serve as a novel anticancer target for bladder transitional cell carcinoma (BTCC).

Highlights

Bladder cancer is the fourth most common cancer and is the eighth leading cause of cancer deaths among men in the world [1]
We focused on the Cullin-RING ligases (CRL), which are the largest family of E3 ubiquitin ligases
Our data showed that Regulator of Cullins-1 (ROC1) protein was weakly expressed in 18 of 24 (75%) normal bladder urothelium, whereas ROC1 protein was highly expressed in bladder cancer tissues (Fig. 1), i.e., among these 112 bladder transitional cell carcinoma (BTCC) tissue specimens, ROC1 protein was moderately or strongly expressed in 29 (25.9%), and 66 (58.9%) samples, respectively

Summary

Introduction

Bladder cancer is the fourth most common cancer and is the eighth leading cause of cancer deaths among men in the world [1]. Bladder transitional cell carcinoma (BTCC) is the most common subtype and accounts for ,90% of all bladder cancers [2]. 20% to 30% of newly diagnosed bladder cancer cases have invaded into the muscle layer (called muscleinvasive transitional cell carcinoma, MI-TCC), while an additional 10% to 30% of nonmuscle-invasive bladder cancers will eventually progress to MI-TCC [3]. The treatment of bladder cancer depends on the depth of the tumor invades into the bladder wall; for MI-TCC patients, cisplatin-based chemotherapy is usually recommended following surgery [2]. The severe toxicity of chemotherapies and relatively low anticancer efficiency confine its widely application in the clinic and majority of MI-TCC will eventually progress and recurrence, leading to poor prognoses of MI-TCC patients [2,4]. Identification of novel effective anticancer targets and agents is of great clinical significance and urgently needed

Methods

Results

Conclusion