The Role Of NOB1in The Development Of Osteosarcoma And Its Mechanism

Osteosarcoma is the most common histological form of primary bone cancer andoften occurs in young people, accounting for5%of childhood cancers and8.9%ofcancer-related deaths in children. The overall incidence is five cases per millionpersons per year. Despite recent improvements in modern multidisciplinary treatmentsincluding chemotherapy and surgery, the5-year survival rate of osteosarcoma patientsremains60-70%. Therefore, we are badly in need of a novel approaches to treatosteosarcoma patients. Recent progress in identifying tumor-associated pathways andnovel gene targeting therapy has led to novel therapeutic strategies and approaches forthe prevention and treatment of osteosarcoma. The current key genes associated withosteosarcoma development including FAS, Ezrin, CXCR4, COX-2, HER2, NOTCH,HES1and so on. Despite the progress, there still lacks an iconic molecule that canaccurately predict progression, metastasis and chemotherapy reactivity ofosteosarcoma.Nin one binding (NOB1) protein is a subunit of the26S proteasome and plays acrucial role in protease function and RNA metabolism. Recent studies have beenincreasingly recognized that abnormal expression of NOB1plays a significant role ina wide variety of tumor types. In human papillary thyroid carcinomas research, NOB1was higher expressed in papillary thyroid carcinomas, compared with normal thyroidand benign thyroid tumour tissue. In human ovarian cancer cells research, knockdownof NOB1expression markedly reduced the proliferative and colonyformation abilityof ovarian cancer cells, and tended to arrest in the G0/G1phase. NOB1deletion alsocaused significant decline in cell proliferation in breast cancer cell lines.Besides,knockdown of NOB1suppressed the proliferation and tumor growth of non-small celllung cancer in vitro and in vivo. However, the importance of NOB1in humanosteosarcoma is largely unknown.In our study, we employed shRNA to knockdown NOB1in osteosarcoma cells, explored the effects of NOB1silencing on cell growth and migration, andscreened the differently expressed genes between Lv-shCon-U2OS cells andLv-shNOB1-U2OS cells using cDNA microarray. In order to investigate the role ofNOB1in osteosarcoma, NOB1expression in human osteosarcoma cell lines wasmeasured using western blot analysis. We employed lentivirus-mediated short hairpinRNA (shRNA） to knockdown NOB1and explored the effects of NOB1silencing oncell growth by MTT, colony formation and cell cycle assays. Cell migration wasobserved with Transwell assay. In addition, the expression levels of E-cadherin andbeta-catenin were examined by qPCR. Moreover, the whole genes mRNA expressionlevels were detected in both control Lv-shCon-U2OS cells and Lv-shNOB1-U2OScells in which NOB1was knocked out using Agilent Human Gene cDNA microarray,and the differently expressed genes were screened and analyzed. Functional analysisindicated that NOB1knockdown strongly inhibited cell growth along with G2/Mphase arresting in human osteosarcoma cells. Moreover, NOB1inhibition decreasedcell migration and increased E-cadherin and beta-catenin expression in U2OS cells.After knockdown of NOB1in U2OS cells, we found792genes have beenup-regulated and1059genes down-regulated. Their encoding proteins were involvedin plasma membrane, calcium ion bindingactivity and signal transduction. Theseresults suggest that NOB1depletion may inhibit osteosarcoma development andprovide for the first time the target potential of NOB1in osteosarcoma treatment.