Xu Wei, Jiang Guo-Jun, Shi Guo-Zhen, Chen Ming-Zhi, Ma Tie-Liang, Tan Yong-Fei
Department of Cardiothoracic Surgery, The Second Affiliated Hospital of Soochow University, Suzhou 215004, China.
Department of Cardiothoracic Surgery, Yixing People's Hospital, Yixing, Wuxi 215004, China.
Transl Cancer Res. 2020 Apr;9(4):2710-2720. doi: 10.21037/tcr.2020.02.61.
Metallothionein 1M (MT1M) functions to regulate cell proliferation and cancer metastasis. This study assessed the effects of MT1M overexpression and mouse double minute 2 homolog (MDM2) knockdown on the regulation of non-small cell lung cancer A549 cell viability, migration, and protein expression and explored the underlying molecular events.
A549 cells were stably infected with lentivirus carrying MT1M cDNA or transiently transfected MDM2 siRNA and/or treated with the p53 inhibitor for the assessment of changes in cell viability, wound healing, Transwell migration, and qRT-PCR and Western blot assays. Luciferase reporter assay was performed to investigate p53 binding to the MT1M promoter.
The data showed that MT1M overexpression inhibited A549 cell viability and migration capacity , whereas the p53 inhibitor reversed the inhibition of A549 cell viability and migration caused by MT1M overexpression as well as the expression of MMP2, MMP9, and MMP14. Furthermore, knockdown of MDM2, an upstream inhibitor of p53 activity, was able to reduce A549 cell viability, migration, and protein expression. Thus, MDM2 knockdown had synergistic effects with MT1M overexpression on the suppression of A549 cell viability, migration, and protein expression.
In conclusion, MDM2 can bind to and phosphorylate p53 protein to inactivate the protein, thereby reducing MT1M expression and leading to tumor cell proliferation and migration.
金属硫蛋白1M（MT1M）具有调节细胞增殖和癌症转移的功能。本研究评估了MT1M过表达和小鼠双微体2同源物（MDM2）敲低对非小细胞肺癌A549细胞活力、迁移及蛋白表达调控的影响，并探究其潜在分子机制。
用携带MT1M cDNA的慢病毒稳定感染A549细胞，或瞬时转染MDM2小干扰RNA（siRNA）和/或用p53抑制剂处理，以评估细胞活力、伤口愈合、Transwell迁移的变化，以及进行定量逆转录聚合酶链反应（qRT-PCR）和蛋白质印迹分析。进行荧光素酶报告基因检测以研究p53与MT1M启动子的结合情况。
数据显示，MT1M过表达抑制了A549细胞活力和迁移能力，而p53抑制剂逆转了MT1M过表达对A549细胞活力和迁移的抑制作用，以及基质金属蛋白酶2（MMP2）、基质金属蛋白酶9（MMP9）和基质金属蛋白酶14（MMP14）的表达。此外，敲低p53活性的上游抑制剂MDM2能够降低A549细胞活力、迁移能力及蛋白表达。因此，敲低MDM2与MT1M过表达在抑制A549细胞活力、迁移及蛋白表达方面具有协同作用。
总之，MDM2可与p53蛋白结合并使其磷酸化，从而使该蛋白失活，进而降低MT1M表达，导致肿瘤细胞增殖和迁移。
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