The TP53 tumor suppressor gene is the most frequently altered gene in tumors and mutant p53 isoforms can acquire oncogenic properties referred to as gain-of-function (GOF). In this study, we used wild-type (A375) and mutant p53 (MeWo) melanoma cell lines to assess the regulation of the mitochondrial antioxidant manganese superoxide dismutase (MnSOD) by mutant p53. The effects of mutant p53 were evaluated by qPCR, immunoblotting, enzyme activity assay, cell proliferation assay, reactive oxygen species (ROS) assay after cellular transfection. We demonstrate that mutant p53 induces MnSOD expression, which is recovered by the ROS scavenger N-acetyl-l-cysteine. This suggests MnSOD induction as a defense mechanism of melanoma cells to counterbalance the pro-oxidant conditions induced by mutant p53. We also demonstrate that mutant p53 induces the expression of Sirtuin3 (SIRT3), a major mitochondrial NAD+-dependent deacetylase, stimulating MnSOD deacetylation and enzymatic activity. Indeed, the restoration of SIRT3 reverses MnSOD activity decrease by mutant p53 knock-down. Finally, MnSOD knock-down further enhances mutant p53-mediated ROS increase, counteracting mutp53-dependent cell hyperproliferation. This indicates that SIRT3 and MnSOD act to maintain ROS levels controlled to promote cell proliferation and survival, providing new therapeutic opportunities to be further considered for clinical studies in cancer patients bearing mutant TP53 gene.

中文翻译：

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p53突变体诱导黑素瘤细胞中SIRT3 / MnSOD轴适度产生ROS。

TP53抑癌基因是肿瘤中最频繁改变的基因，突变的p53亚型可以获得致癌特性，称为功能获得（GOF）。在这项研究中，我们使用野生型（A375）和突变型p53（MeWo）黑色素瘤细胞系来评估突变型p53对线粒体抗氧化剂锰超氧化物歧化酶（MnSOD）的调节。通过qPCR，免疫印迹，酶活性测定，细胞增殖测定，细胞转染后的活性氧（ROS）测定来评估突变体p53的作用。我们证明突变体p53诱导MnSOD表达，这是由ROS清道夫N-乙酰-1-半胱氨酸回收的。这表明MnSOD诱导作为黑色素瘤细胞的防御机制，以抵消突变体p53诱导的促氧化条件。我们还证明，突变体p53诱导Sirtuin3（SIRT3）的表达，主要的线粒体NAD +依赖性脱乙酰基酶，刺激MnSOD脱乙酰基化和酶活性。实际上，SIRT3的恢复逆转了通过突变p53敲低而降低的MnSOD活性。最后，MnSOD组合式进一步增强了突变体p53介导的ROS的增加，抵消了mutp53依赖性细胞的过度增殖。这表明SIRT3和MnSOD的作用是维持受控的ROS水平，从而促进细胞增殖和存活，从而为具有突变型TP53基因的癌症患者的临床研究提供了新的治疗机会。SIRT3的恢复逆转了MnSOD活性因突变体p53敲低而降低的趋势。最后，MnSOD组合式进一步增强了突变体p53介导的ROS的增加，抵消了mutp53依赖性细胞的过度增殖。这表明SIRT3和MnSOD的作用是维持受控的ROS水平，从而促进细胞增殖和存活，从而为具有突变型TP53基因的癌症患者的临床研究提供了新的治疗机会。SIRT3的恢复逆转了MnSOD活性因突变体p53敲低而降低的趋势。最后，MnSOD组合式进一步增强了突变体p53介导的ROS的增加，抵消了mutp53依赖性细胞的过度增殖。这表明SIRT3和MnSOD的作用是维持受控的ROS水平，从而促进细胞增殖和存活，从而为具有突变型TP53基因的癌症患者的临床研究提供了新的治疗机会。