Department of Cancer Biology and Genetics, The Ohio State University Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210;
Department of Biosciences, John van Geest Cancer Research Centre, School of Science and Technology, Nottingham Trent University, NG11 8NS Nottingham, United Kingdom.
Proc Natl Acad Sci U S A. 2020 Nov 17;117(46):28806-28815. doi: 10.1073/pnas.2017830117. Epub 2020 Nov 2.
Akt activation up-regulates the intracellular levels of reactive oxygen species (ROS) by inhibiting ROS scavenging. Of the Akt isoforms, Akt3 has also been shown to up-regulate ROS by promoting mitochondrial biogenesis. Here, we employ a set of isogenic cell lines that express different Akt isoforms, to show that the most robust inducer of ROS is Akt3. As a result, Akt3-expressing cells activate the DNA damage response pathway, express high levels of p53 and its direct transcriptional target miR-34, and exhibit a proliferation defect, which is rescued by the antioxidant -acetylcysteine. The importance of the DNA damage response in the inhibition of cell proliferation by Akt3 was confirmed by Akt3 overexpression in and / mouse embryonic fibroblasts (MEFs), which failed to inhibit cell proliferation, despite the induction of high levels of ROS. The induction of ROS by Akt3 is due to the phosphorylation of the NADPH oxidase subunit p47, which results in NADPH oxidase activation. Expression of Akt3 in MEFs failed to induce ROS and to inhibit cell proliferation. Notably, the proliferation defect was rescued by wild-type p47, but not by the phosphorylation site mutant of p47 In agreement with these observations, Akt3 up-regulates p53 in human cancer cell lines, and the expression of Akt3 positively correlates with the levels of p53 in a variety of human tumors. More important, alterations correlate with a higher frequency of mutation of , suggesting that tumor cells may adapt to high levels of Akt3, by inactivating the DNA damage response.
Akt 的激活通过抑制 ROS 清除来上调细胞内活性氧 (ROS) 的水平。在 Akt 同工型中，Akt3 还通过促进线粒体生物发生来上调 ROS。在这里，我们使用一组表达不同 Akt 同工型的同基因细胞系，表明 Akt3 是最能强烈诱导 ROS 的同工型。结果，表达 Akt3 的细胞激活了 DNA 损伤反应途径，表达高水平的 p53 及其直接转录靶标 miR-34，并表现出增殖缺陷，抗氧化剂 -乙酰半胱氨酸可挽救该缺陷。Akt3 在 和 / 鼠胚胎成纤维细胞 (MEF) 中的过表达未能抑制细胞增殖，尽管诱导了高水平的 ROS，但证实了 DNA 损伤反应在 Akt3 抑制细胞增殖中的重要性。Akt3 诱导 ROS 的原因是 NADPH 氧化酶亚基 p47 的磷酸化，导致 NADPH 氧化酶激活。在 MEF 中表达 Akt3 未能诱导 ROS 和抑制细胞增殖。值得注意的是，野生型 p47 可挽救增殖缺陷，但磷酸化位点突变型 p47 则不能。与这些观察结果一致，Akt3 在人癌细胞系中上调 p53，并且 Akt3 的表达与各种人肿瘤中 p53 的水平呈正相关。更重要的是， 改变与 的突变频率更高相关，这表明肿瘤细胞可能通过使 DNA 损伤反应失活来适应高水平的 Akt3。
