新型癌基因 CUG2 诱导的 EGFR 表达增加通过 Stat1-HDAC4 信号通路赋予多柔比星耐药性。
Increased EGFR expression induced by a novel oncogene, CUG2, confers resistance to doxorubicin through Stat1-HDAC4 signaling.
机构信息
BK21+, Department of Cogno-Mechatronics Engineering, Pusan National University, Busan, 46241, Republic of Korea.
Department of Nanoenergy Engineering, Pusan National University, Busan, 46241, Republic of Korea.
出版信息
Cell Oncol (Dordr). 2017 Dec;40(6):549-561. doi: 10.1007/s13402-017-0343-7. Epub 2017 Aug 3.
BACKGROUND
Previously, it has been found that the cancer upregulated gene 2 (CUG2) and the epidermal growth factor receptor (EGFR) both contribute to drug resistance of cancer cells. Here, we explored whether CUG2 may exert its anticancer drug resistance by increasing the expression of EGFR.
METHODS
EGFR expression was assessed using Western blotting, immunofluorescence and capacitance assays in A549 lung cancer and immortalized bronchial BEAS-2B cells, respectively, stably transfected with a CUG2 expression vector (A549-CUG2; BEAS-CUG2) or an empty control vector (A549-Vec; BEAS-Vec). After siRNA-mediated EGFR, Stat1 and HDAC4 silencing, antioxidant and multidrug resistance protein and mRNA levels were assessed using Western blotting and RT-PCR. In addition, the respective cells were treated with doxorubicin after which apoptosis and reactive oxygen species (ROS) levels were measured. Stat1 acetylation was assessed by immunoprecipitation.
RESULTS
We found that exogenous CUG2 overexpression induced EGFR upregulation in A549 and BEAS-2B cells, whereas EGFR silencing sensitized these cells to doxorubicin-induced apoptosis. In addition, we found that exogenous CUG2 overexpression reduced the formation of ROS during doxorubicin treatment by enhancing the expression of antioxidant and multidrug resistant proteins such as MnSOD, Foxo1, Foxo4, MRP2 and BCRP, whereas EGFR silencing congruently increased the levels of ROS by decreasing the expression of these proteins. We also found that EGFR silencing and its concomitant Akt, ERK, JNK and p38 MAPK inhibition resulted in a decreased Stat1 phosphorylation and, thus, a decreased activation. Since also acetylation can affect Stat1 activation via a phospho-acetyl switch, HDAC inhibition may sensitize cells to doxorubicin-induced apoptosis. Interestingly, we found that exogenous CUG2 overexpression upregulated HDAC4, but not HDAC2 or HDAC3. Conversely, we found that HDAC4 silencing sensitized the cells to doxorubicin resistance by decreasing Stat1 phosphorylation and EGFR expression, thus indicating an interplay between HDAC4, Stat1 and EGFR.
CONCLUSION
Taken together, we conclude that CUG2-induced EGFR upregulation confers doxorubicin resistance to lung (cancer) cells through Stat1-HDAC4 signaling.
背景
先前的研究发现，癌上调基因 2（CUG2）和表皮生长因子受体（EGFR）均有助于癌细胞的耐药性。在这里，我们探讨了 CUG2 是否可以通过增加 EGFR 的表达来发挥其抗癌药物耐药性。
方法
分别使用 Western blot、免疫荧光和电容测定法评估稳定转染 CUG2 表达载体（A549-CUG2；BEAS-CUG2）或空载体对照（A549-Vec；BEAS-Vec）的 A549 肺癌和永生化支气管 BEAS-2B 细胞中的 EGFR 表达。用 siRNA 介导 EGFR、Stat1 和 HDAC4 沉默后，使用 Western blot 和 RT-PCR 评估抗氧化剂和多药耐药蛋白和 mRNA 水平。此外，在用阿霉素处理后，分别测量细胞的凋亡和活性氧（ROS）水平。通过免疫沉淀评估 Stat1 乙酰化。
结果
我们发现，外源性 CUG2 过表达诱导 A549 和 BEAS-2B 细胞中 EGFR 的上调，而 EGFR 沉默则使这些细胞对阿霉素诱导的凋亡敏感。此外，我们发现，外源性 CUG2 过表达通过增强抗氧化剂和多药耐药蛋白（如 MnSOD、Foxo1、Foxo4、MRP2 和 BCRP）的表达，减少阿霉素处理过程中 ROS 的形成，而 EGFR 沉默则通过降低这些蛋白的表达，一致增加 ROS 的水平。我们还发现，EGFR 沉默及其伴随的 Akt、ERK、JNK 和 p38 MAPK 抑制导致 Stat1 磷酸化减少，从而导致激活减少。由于磷酸-乙酰化开关也可以影响 Stat1 的激活，因此 HDAC 抑制可能使细胞对阿霉素诱导的凋亡敏感。有趣的是，我们发现外源性 CUG2 过表达上调了 HDAC4，但没有上调 HDAC2 或 HDAC3。相反，我们发现 HDAC4 沉默通过降低 Stat1 磷酸化和 EGFR 表达使细胞对阿霉素耐药，这表明 HDAC4、Stat1 和 EGFR 之间存在相互作用。
结论
综上所述，我们得出结论，CUG2 诱导的 EGFR 上调通过 Stat1-HDAC4 信号赋予肺癌（癌）细胞阿霉素耐药性。
Zotero 插件