摘要
目的探讨精氨酸甲基转移酶1(CARM1)调控铁死亡在肺癌细胞恶性生物学中的分子机制。方法通过免疫印迹检测正常人支气管上皮细胞(HBE4)和肺癌系中(A549、H1299、H1640、HCC827)中CARM1的表达情况。将CARM1序列或载体对照(Vector)以及针对CARM1(shCARM1)、Notch同源物2(shNotch2)的shRNA序列或阴性对照shRNA(shNC)转染到肺癌细胞中。分别通过CCK⁃8试验、Transwell试验测定细胞增殖、迁移和侵袭。利用RIP⁃PCR和MeRIP⁃qPCR分析探讨了CARM1的作用机制。采用经典的铁死亡诱导剂Erastin处理肺癌细胞以确定CARM1是否能影响细胞对铁死亡的敏感性。结果与人正常气道上皮细胞HBE4相比,CARM1在肺癌系中(A549、H1299、H1640、HCC827)显著上调(P<0.05)。与Vector组相比,CARM1过表达组A549细胞增殖、迁移和侵袭能力显著增加(P<0.001),而shCARM1组HCC827细胞的增殖、迁移和侵袭能力显著低于shNC组(P<0.001)。MeRIP⁃qPCR分析显示当CARM1过表达时Notch2 mRNA的m6A丰度显著增加(P<0.05)。RIP分析显示CARM1过表达显著促进CARM1和Notch2 mRNA之间的结合(P<0.05)。Notch2敲低减弱了CARM1上调的A549细胞的增殖、侵袭和迁移能力(P<0.001)。结论CARM1在肺癌中显著升高,并通过激活Notch2发挥其致癌功能。此外,CARM1可以通过稳定Notch2使肺癌细胞对铁死亡不敏感。
Objective To explore the molecular mechanism of arginine methyltransferase 1(CARM1)reg⁃ulating ferroptosis in malignant biology of lung cancer cells.Methods The expression of CARM1 in normal human bronchial epithelial cells(HBE4)and lung cancer lines(A549,H1299,H1640,HCC827)was detected by west⁃ern blot.CARM1 sequence or Vector and shRNA sequence against CARM1(shCARM1)and Notch homologue 2(shNotch2)or negative control shRNA(shNC)were transfected into lung cancer cells.Cell proliferation,migration and invasion were measured by CCK⁃8 test and Transwell test respectively.RIP⁃PCR and MeRIP⁃qPCR analysis were used to explore the mechanism of CARM1.Erastin,a classical ferroptosis inducer,was used to treat lung can⁃cer cells to determine whether CARM1 could affect the sensitivity of cells to ferroptosis.Results Compared with human normal airway epithelial cell HBE4,CARM1 was significantly over⁃expressed in lung cancer lines(A549,H1299,H1640,HCC827)(P<0.05).Compared with Vector group,the proliferation,migration and invasion ability of A549 cells in CARM1 group was significantly increased(P<0.001),while that of HCC827 cells in shCARM1 group was significantly lower than that in shNC group(P<0.001).MeRIP⁃qPCR analysis showed that m6A abundance of Notch2 mRNA increased significantly when CARM1 was overexpressed(P<0.05).RIP analy⁃sis showed that CARM1 overexpression significantly promoted the binding between CARM1 and Notch2 mRNA(P<0.05).Notch2 knockdown weakened the proliferation,invasion and migration of A549 cells up⁃regulated by CARM1(P<0.001).Conclusion CARM1 is significantly elevated in lung cancer,and plays its carcinogenic role by activating Notch2.In addition,CARM1 can make lung cancer cells insensitive to ferroptosis by stabilizing Notch2.
作者
卢天龙
杨启英
杨世闻
宋小龙
LU Tian-long;YANG Qiying;YANG Shiwen;SONG Xiaolong(Department of Clinical Laboratory,Qinghai University Affili-ated Hospital,Xining 810000,China)
出处
《实用医学杂志》
CAS
北大核心
2023年第9期1098-1104,共7页
The Journal of Practical Medicine
基金
青海省自然科学基金项目(编号:2019⁃ZY⁃2631)。
