TRIM3通过调控PIAS1-STAT1轴抑制胶质瘤发生发展  

TRIM3 inhibits glioblastoma progression by regulating the PIAS1-STAT1 axis

作　　者：陈昱宁 焦建同 郭卉 计巍 何其晟 邵君飞 CHEN Yuning;JIAO Jiantong;GUO Hui;JI Wei;HE Qisheng;SHAO Junfei(Department of Neurosurgery,Wuxi People s Hospital,Wuxi 214023,China)

机构地区：[1]无锡市人民医院神经外科,南京医科大学附属无锡人民医院,南京医科大学无锡医学中心,无锡214023

出　　处：《临床神经外科杂志》2025年第2期149-157,165,共10页Journal of Clinical Neurosurgery

基　　金：江苏省卫生健康委2022年度医学科研重点项目(ZD2022038)。

摘　　要：目的 探究TRIM3通过PIAS1-STAT1在胶质母细胞瘤(GBM)形成与发展中发挥的作用及其分子机制。方法 构建TRIM3的过表达慢病毒与shRNA转染的GBM细胞系U251与U87细胞,同时设置相应的对照组CON335和CON313。使用FLAG-TRIM3质粒转染GBM细胞与293T细胞,构建过表达细胞系。活细胞计数(CCK-8)、平板克隆实验、Transwell小室及流式细胞术对胶质瘤细胞的增殖活性、成团、迁移能力以及凋亡情况进行分析。Western blot和免疫共沉淀实验(Co-IP)用于检测TRIM3与PIAS1蛋白互作情况以及具体分子机制。实时定量荧光聚合酶链反应(PCR)检测TRIM3和PIAS1的mRNA表达。双荧光素酶报告基因试验检测TRIM3对STAT1的活性影响。裸鼠颅内成瘤模型证明TRIM3在体内对胶质瘤形成的影响。结果 与正常脑组织细胞相比,TRIM3的表达在GBM组织中明显降低,并且TRIM3的表达水平随着胶质瘤WHO病理分级增高而降低。与对照组相比,在过表达了TRIM3之后,胶质瘤细胞的增殖、成团与迁移能力明显降低,并且可诱导GBM细胞的凋亡。Western blot结果显示,TRIM3在GBM细胞中过表达会导致PIAS1蛋白水平的降低,且浓度越高趋势越明显。实时定量荧光PCR结果提示TRIM3对PIAS1和STAT1的mRNA水平无影响。双荧光素酶报告基因试验证明TRIM3提高STAT1的转录活性。此外,TRIM3通过促进PIAS1在K48位点的多聚泛素化介导其降解,从而解除PIAS1对STAT1转录效应的抑制。在裸鼠体内同样证明与对照组相比,TRIM3过表达组小鼠颅内成瘤速度与大小明显下降。结论 TRIM3拮抗PIAS1-TRIM3通路,抑制胶质瘤发生发展,为肿瘤治疗提供新的靶点。Objective To explore the role and molecular mechanisms of TRIM3 in the occurrence and development of human glioblastoma through PIAS1-STAT1.Methods Lentiviral overexpression of TRIM3 and shRNA-transfected GBM cell lines U251 and U87 were constructed,with corresponding control groups CON335 and CON313.The FLAG-TRIM3 plasmid was used to transfect GBM cells and 293T cells,to construct overexpression cell lines.Living cell counting(CCK-8),plate cloning experiments,Transwell chambers,and flow cytometry were used to analyze the proliferative activity,clumping,migration ability,and apoptosis of glioma cells.Western blot and co-immunoprecipitation(Co-IP) experiments were conducted to detect the interaction between TRIM3 and PIAS1 proteins,as well as the specific molecular mechanisms.Real-time quantitative fluorescent polymerase chain reaction(PCR) was used to detect the mRNA expression of TRIM3 and PIAS1.Dual-luciferase reporter gene assays were employed to test the impact of TRIM3 on STAT1 activity.Nude mouse intracranial tumor models were used to demonstrate the impact of TRIM3 on glioma formation in vivo.Results Compared to normal brain tissue cells,the expression of TRIM3 was significantly reduced in GBM tissues.The expression level of TRIM3 decreased with the WHO grades.Overexpression ofTRIM3 in glioma cells resulted in a significant attenuation of their proliferative,clumping,and migratory capacities,and induced apoptosis in GBM cells.Western blot analysis revealed that overexpression of TRIM3 in GBM cells was associated with a concentrationdependent reduction in PIAS1 protein levels.Real-time quantitative PCR data indicated thatTRIM3 overexpression had no influence on the mRNA levels of PIAS1 and STAT1.Dual-luciferase reporter assays confirmed that TRIM3 enhances the transcriptional activity of STAT1.Moreover,TRIM3 mediated the degradation of PIAS1 by promoting its poly-ubiquitination at the K48 residue,thus alleviating the suppressive effect of PIAS1 on STAT1 transcription.In vivo studies using a nude mouse model

关 键 词：胶质母细胞瘤 TRIM3 PIAS1 STAT1 泛素化 

分 类 号：R739.41[医药卫生—肿瘤]