摘要
目的 探讨BTB与CNC同源蛋白1(BACH1)通过调节MYC相关因子X二聚化蛋白1(MXD1)基因对套细胞淋巴瘤(MCL)发生发展的影响。方法 采用靶向剪切及转座酶技术(CUT&Tag)检测BACH1下游靶基因,并通过序列比对分析寻找BACH1与MXD1的结合位点。通过双荧光素酶报告基因系统验证BACH1对MXD1的调控作用,并采用慢病毒技术构建BACH1过表达及敲低的稳转MCL细胞株,采用蛋白质印迹法(Western blot)检测MXD1蛋白表达水平。应用GSE16411数据集分析MCL细胞和正常B细胞中MXD1 m RNA相对表达量。应用GSE132929及GSE21452数据集筛选MXD1共表达基因,对MXD1共表达基因进行基因本体(GO)和京都基因与基因组百科全书(KEGG)信号通路富集分析。采用噻唑蓝(MTT)法检测MXD1正相关基因应激诱导磷蛋白1同源物含U-框蛋白1(STUB1)的特异性激活剂YL109处理后MCL细胞的活力。结果 BACH1能够靶向结合MXD1并抑制MXD1基因的启动子活性。BACH1敲低后MXD1蛋白表达水平升高,而BACH1过表达后MXD1蛋白表达水平降低。通过GSE16411数据集分析发现,MCL细胞中MXD1 mRNA相对表达量明显低于正常B细胞,差异有统计学意义(P﹤0.01)。在GSE132929数据集中筛选出2222个MXD1共表达基因,在GSE21452数据集中筛选出503个MXD1共表达基因。GO分析显示,MXD1共表达基因主要富集于信号转导、蛋白质磷酸化、细胞葡萄糖醛酸化、类黄酮葡萄糖醛酸化等代谢相关的生物学过程;KEGG信号通路分析显示,MXD1共表达基因主要富集于肝脏发育的代谢途径、蛋白质的消化和吸收、Janus激酶(JAK)/信号转导及转录激活因子(STAT)信号通路等。采用YL109处理MCL细胞后可显著降低细胞存活率。结论 BACH1通过结合MXD1近端启动子负向调控MXD1的表达水平,且BACH1介导的MXD1表达改变很可能通过调控代谢过程参与MCL的发生发展。
Objective To explore the effect of BTB domain and CNC homolog 1(BACH1)on the occurrence and de-velopment of mantle cell lymphoma(MCL)by regulating the MYC associated factor X dimerization protein 1(MXD1)gene.Method Cleavage under targets and tagmentation(CUT&Tag)was used to detect the downstream target genes of BACH1,and sequence alignment was used to identify the binding sites of BACH1 and MXD1.Dual luciferase reporter gene system was used to verify the regulation of BACH1 on MXD1.BACH1 overexpression and knockdown stable MCL cell lines were generated by lentivirus-mediated system,and the expression levels of MXD1 protein in these cells were de-tected by Western blot.The GSE16411 dataset was used to analyze the relative expression of MXD1 mRNA in MCL cells and normal B cells.GSE132929 and GSE21452 datasets were used to screen out MXD1 co-expressed genes,and Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)signaling pathway enrichment analysis were performed for MXD1 co-expressed genes.Methyl thiazolyl tetrazolium(MTT)assay was used to analyze the cell viability after treatment of YL109,a specific activator of MXD1 positive related gene stress-induced-phosphoprotein 1 homology and U-box containing protein 1(STUB1).Result BACH1 targeted binding MXD1 and inhibited the activity of MXD1 promoter.The expression level of MXD1 protein increased after BACH1 knockdown,while the expression level of MXD1 protein decreased after BACH1 overexpression.The analysis of GSE16411 dataset showed that the relative expression lev-el of MXD1 mRNA was significantly lower than that of normal B cells,and the difference was statistically significant(P<0.01).A total of 2222 MXD1 co-expressed genes were screened in GSE132929 dataset and 503 MXD1 co-expressed genes were screened in GSE21452 dataset.GO analysis showed that MXD1 co-expressed genes were mainly enriched in metabolic related biological processes such as signal transduction,protein phosphorylation,cellular glucosylation,and fla-vonoid glucosylation.KEGG signaling
作者
张紫婷
张寒
ZHANG Ziting;ZHANG Han(Institute of Medical Biology,Chinese Academy of Medical Sciences&Peking Union Medical College,Kunming 650118,Yunnan,China)
出处
《癌症进展》
2024年第6期610-618,共9页
Oncology Progress
基金
云南省卫生健康委员会医学学科带头人培养计划(D-2019027)
云南省科技厅科技计划项目(202301AS070067)。
