Niu Yichao, Xia Ye, Wang Jingyun, Shi Xiaofei
Department of Gynecology, Shanghai First Maternity and Infant Hospital, Tongji University School of Medicine, Shanghai 200126, P.R. China.
Mol Med Rep. 2017 Apr;15(4):2083-2089. doi: 10.3892/mmr.2017.6244. Epub 2017 Feb 24.
O-GlcNAcylation is a dynamic and reversible post-translational modification associated with the regulation of multiple cellular functions. The addition and removal of O‑Linked β-N-acetylglucosamine (O‑GlcNAc) on target proteins is catalyzed by O‑GlcNAc transferase (OGT) and O‑GlcNAcase (OGA), respectively. Accumulating evidence suggests that O-GlcNAcylation is associated with the malignancy of several types of human cancer. To investigate the effect of O-GlcNAcylation on ovarian cancer phenotypes, global O‑GlcNAc levels were decreased by OGT silencing through RNA interference and increased by inhibiting OGA activity with Thiamet‑G. Transwell assay results demonstrated that OGT silencing inhibited the migration and invasion of SKOV3 and 59M ovarian cells in vitro, while Thiamet‑G treatment promoted migration and invasion. Furthermore, a pull‑down assay and western blot analysis demonstrated that Thiamet-G treatment enhanced RhoA activity and the phosphorylation of the Rho‑associated protein kinase (ROCK) substrate, myosin light chain (MLC), while OGT silencing attenuated RhoA activity and MLC phosphorylation. In addition, RhoA silencing via RNA interference and inhibition of ROCK activity with Y‑27632 prevented Thiamet‑G‑induced increases in cell migration and invasion. These data suggest that O‑GlcNAcylation augments the motility of ovarian cancer cells via the RhoA/ROCK/MLC signaling pathway. Therefore, O‑GlcNAcylation may be a potential target for the diagnosis and treatment of ovarian cancer.
O-连接的N-乙酰葡糖胺（O-GlcNAc）化是一种与多种细胞功能调节相关的动态可逆的翻译后修饰。靶蛋白上O-连接的β-N-乙酰葡糖胺（O-GlcNAc）的添加和去除分别由O-GlcNAc转移酶（OGT）和O-GlcNAcase（OGA）催化。越来越多的证据表明，O-GlcNAc化与几种类型的人类癌症的恶性程度有关。为了研究O-GlcNAc化对卵巢癌表型的影响，通过RNA干扰使OGT沉默来降低整体O-GlcNAc水平，并使用噻美汀-G抑制OGA活性来提高该水平。Transwell实验结果表明，OGT沉默在体外抑制了SKOV3和59M卵巢癌细胞的迁移和侵袭，而噻美汀-G处理则促进了迁移和侵袭。此外，下拉实验和蛋白质印迹分析表明，噻美汀-G处理增强了RhoA活性以及Rho相关蛋白激酶（ROCK）底物肌球蛋白轻链（MLC）的磷酸化，而OGT沉默则减弱了RhoA活性和MLC磷酸化。此外，通过RNA干扰使RhoA沉默以及用Y-27632抑制ROCK活性可阻止噻美汀-G诱导的细胞迁移和侵袭增加。这些数据表明，O-GlcNAc化通过RhoA/ROCK/MLC信号通路增强卵巢癌细胞的运动性。因此，O-GlcNAc化可能是卵巢癌诊断和治疗的一个潜在靶点。
