Login to Access Video or Poster Abstract: MP48-05
Sources of Funding: National Natural Science Foundation of China (No.81172419)

Introduction

The coiled coil is a superhelical structural protein motif involved in a diverse array of biological functions, and the abnormal expression of the coiled-coil domain containing proteins has a direct link with the phenotype of tumor cell migration, invasion and metastasis. Here, we first reported the oncogenic roles of Coiled-coil domain-containing protein 34 (CCDC34), and investigated its biological functions in bladder carcinogenesis.

Methods

Immunohistochemical staining and western blot were used to detect CCDC34 expression in bladder cancers specimens and cell lines. Lentivirus-mediated RNA interference and overexpression strategies were used to assess the effects of CCDC34 expression on various malignant phenotypes. The biological functions of CCDC34 knockdown on cells (T24 and 5637) were investigated by examining cell proliferation using a high content screening assay (HCS), BrdU incorporation assay and colony formation assay, cell migration by in vitro wound healing assay, cell invasion by Transwell invasion assay, as well as cell cycle distribution and apoptosis by flow cytometry. The expressions of Bcl-2, c-Raf, c-Jun, N-cadherin and E-cadherin as well as the phosphorylation of MEK, ERK1/2 and AKT were also measured using Western blot. We further investigated the effect of therapeutic siRNA targeting CCDC34 on T24 xenograft tumor growth in nude mice.

Results

CCDC34 was up-regulated in human bladder cancer tissues and cell lines. CCDC34 was distributed mainly in the cytoplasm, and its expression was closely correlated with histological type, tumor grade and pathologic stage (n=87, P<0.05). Besides, Western blot confirmed that CCDC34 was expressed at higher level in human bladder cancer tissues compared with their paraneoplastic normal bladder tissues (n=18, P=0.012). Knockdown of CCDC34 significantly suppressed bladder cancer cells proliferation, migration and invasion (P<0.01), and induced cell cycle arrest at G2/M phase and increased apoptosis in vitro (P<0.01). Moreover, CCDC34 knockdown decreased phosphorylation of MEK, ERK1/2 and AKT, and the expressions of c-Raf, c-Jun and Bcl-2; while CCDC34 overexpression in T24 cells promoted cell migration and invasion with increased EMT, and treatment with MAPK/ERK1/2 and PI3K/AKT inhibitors blocked the phosphorylation of ERK1/2 and AKT, respectively. In addition, knockdown of CCDC34 suppressed bladder cancer cell growth in nude mice.

Conclusions

Our findings revealed for the first time a potential oncogenic role for CCDC34 in bladder carcinoma pathogenesis, and activation of MAPK/ERK1/2 and PI3K/AKT signaling pathways was required for CCDC34 modulation of bladder cancer cell proliferation, migration and invasion. CCDC34 may serve as a biomarker or even a therapeutic target for bladder carcinoma.

Funding

National Natural Science Foundation of China (No.81172419)