Department of Orthopedics, Taizhou First People's Hospital, Taizhou, 318020, Zhejiang, China.
Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.
Biochem Biophys Res Commun. 2019 Jul 5;514(4):1172-1178. doi: 10.1016/j.bbrc.2019.04.188. Epub 2019 May 15.
Osteosarcoma is a highly malignant and aggressive bone tumor. Its occurrence and development involve many factors and multiple signaling pathways. Some studies have shown that circular RNAs (circRNAs) play important roles in the development of various tumors. This research showed that circ_ORC2 was generally up-regulated in various osteosarcoma cell lines, and mainly distributed in the cytoplasm. Circ_ORC2 had the binding site of miR-19a, and its expression was positively correlated with miR-19a expression. RIP experiments showed that circ_ORC2 could bind to Ago2 protein. RNA pull-down using biotinylated circ_ORC2 or miR-19a showed that circ_ORC2 could directly interact with miR-19a, and dual luciferase reporter gene assay also confirmed that miR-19a could bind to circ_ORC2. After circ_ORC2 knockdown, miR-19a expression was down-regulated, but the downstream target gene PTEN expression was up-regulated, and the phosphorylation level of Akt was reduced, which indicated that circ_ORC2 enhanced the inhibition of miR-19a on PTEN expression by combining miR-19a. Further functional experiments showed that after circ_ORC2 knockdown, cell proliferation and invasion decreased, while the apoptosis level increased. When co-transfected with circ_ORC2 siRNA and miR-19a mimics or PTEN siRNA, the above cell biological behaviors did not change significantly. Therefore, circ_ORC2 binds with miR-19a and enhances its expression, thereby inhibiting downstream PTEN expression and activating Akt pathway to promote osteosarcoma cell growth and invasion. These findings enrich the circRNA molecular regulation mechanism, and provide more reference ideas for the research and application of circRNAs in tumors and other diseases.
骨肉瘤是一种高度恶性和侵袭性的骨肿瘤。其发生和发展涉及许多因素和多个信号通路。一些研究表明，环状 RNA（circRNA）在各种肿瘤的发展中发挥着重要作用。这项研究表明，circ_ORC2 在各种骨肉瘤细胞系中普遍上调，主要分布在细胞质中。circ_ORC2 具有 miR-19a 的结合位点，其表达与 miR-19a 表达呈正相关。RIP 实验表明，circ_ORC2 可以与 Ago2 蛋白结合。用生物素标记的 circ_ORC2 或 miR-19a 进行 RNA 下拉实验表明，circ_ORC2 可以直接与 miR-19a 相互作用，双荧光素酶报告基因实验也证实了 miR-19a 可以与 circ_ORC2 结合。circ_ORC2 敲低后，miR-19a 表达下调，但下游靶基因 PTEN 表达上调，Akt 磷酸化水平降低，表明 circ_ORC2 通过结合 miR-19a 增强了对 PTEN 表达的抑制作用。进一步的功能实验表明，circ_ORC2 敲低后，细胞增殖和侵袭减少，而凋亡水平增加。当共转染 circ_ORC2 siRNA 和 miR-19a 模拟物或 PTEN siRNA 时，上述细胞生物学行为没有明显变化。因此，circ_ORC2 与 miR-19a 结合并增强其表达，从而抑制下游 PTEN 表达并激活 Akt 通路，促进骨肉瘤细胞的生长和侵袭。这些发现丰富了 circRNA 分子调控机制，为 circRNA 在肿瘤等疾病中的研究和应用提供了更多的参考思路。
