Background

Osteosarcoma is the most prevalent primary malignant bone tumor containing mesenchymal cells with poor prognosis. Being a hot spot of anti-tumor therapy researches, AKT/mammalian target of rapamycin (mTOR) signaling pathway could affect various cellular processes including transcription, protein synthesis, apoptosis, autophagy and growth.

Materials and methods

The levels of RNA and protein were detected by quantitative real-time polymerase chain reaction (q-PCR) and western blot analyses respectively. Functional assays were carried out to analyze the malignant phenotypes of osteosarcoma cells. RNA-binding protein immunoprecipitation (RIP), Co-immunoprecipitation (Co-IP), RNA pulldown, luciferase reporter and in vitro kinase assays were conducted to uncover the specific mechanism of microRNA-451a (miR-451a) in osteosarcoma cells.

Results

Functionally, miR-451a represses the malignant progression of osteosarcoma. Mechanically, miR-451a could curb the AKT/mTOR pathway via 3-phosphoinositide dependent protein kinase 1 (PDPK1)-mediated phosphorylation modification. After the certification that YTH domain containing 1 (YTHDC1) regulates the m6A phosphorylation modification of PDPK1 mRNA, we further proved that miR-451a-mediated YTHDC1 stabilizes PDPK1 mRNA via m6A-dependent regulation.

Conclusion

This study demonstrated that miR-451a regulates YTHDC1-mediated m6A methylation to activate the AKT/mTOR pathway, stimulating the malignancy of osteosarcoma.

中文翻译：

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MiR-451a 通过调节 YTHDC1 介导的 m6A 甲基化激活 AKT/mTOR 信号通路促进骨肉瘤中的细胞生长、迁移和 EMT

背景

骨肉瘤是最常见的原发性恶性骨肿瘤，含有间充质细胞，预后较差。作为抗肿瘤治疗研究的热点，AKT/哺乳动物雷帕霉素靶蛋白（mTOR）信号通路可以影响多种细胞过程，包括转录、蛋白质合成、细胞凋亡、自噬和生长。

材料和方法

分别通过定量实时聚合酶链反应（q-PCR）和蛋白质印迹分析检测RNA和蛋白质的水平。进行功能测定以分析骨肉瘤细胞的恶性表型。进行了 RNA 结合蛋白免疫沉淀 (RIP)、免疫共沉淀 (Co-IP)、RNA pulldown、荧光素酶报告基因和体外激酶测定，以揭示 microRNA-451a (miR-451a) 在骨肉瘤细胞中的具体机制。

结果

在功能上，miR-451a 抑制骨肉瘤的恶性进展。机械地，miR-451a 可以通过 3-磷酸肌醇依赖性蛋白激酶 1 (PDPK1) 介导的磷酸化修饰来抑制 AKT/mTOR 通路。在证实含有 1 的 YTH 结构域（YTHDC1）调节 PDPK1 mRNA 的 m6A 磷酸化修饰后，我们进一步证明 miR-451a 介导的 YTHDC1 通过 m6A 依赖性调节稳定 PDPK1 mRNA。

结论

本研究表明 miR-451a 调节 YTHDC1 介导的 m6A 甲基化以激活 AKT/mTOR 通路，刺激骨肉瘤的恶性。