【摘要】 目的检测食管胃结合部腺癌(AEG)患者基因突变情况,分析内披蛋白(IVL)突变对肿瘤细胞生物学功能的影响。方法将22例山西地区食管胃结合部腺癌患者的癌组织和癌旁组织进行全外显子组测序,分析IVL的突变情况并筛选IVL突变位点;整理汇总TCGA数据库中食管癌IVL的突变情况,进行GO功能注释和KEGG富集分析。构建IVL过表达质粒,转染食管癌细胞系KYSE150,测定细胞的增殖和迁移能力。体外合成不同突变位点的IVL寡肽,通过体外活性测试分析寡肽与转谷氨酰胺酶1(TGase1)的结合能力。结果测序结果发现在食管胃结合部腺癌中IVL有5处突变;GO分析显示IVL突变与上皮细胞分化、细胞分化调控的活性等生物学过程有关;KEGG信号通路分析显示IVL突变与cAMP信号通路等途径有关。过表达IVL可抑制食管癌细胞的增殖和迁移能力(P<0.05)。突变型IVL寡肽与TGase1的结合能力弱于野生型寡肽(P<0.05)。结论 IVL重复基序第7位氨基酸的突变减弱IVL与TGase1的结合,影响多种与肿瘤相关的生物学过程和信号通路。更多还原

【Abstract】 Objective To detect the gene mutations in patients with Adenocarcinoma at esophagogastric junction(AEG) and to analyze the effect of Involucrin(IVL) mutation on the biological function of tumor cells. MethodsWhole exome sequencing was performed on cancerous and paired adjacent normal tissues from 22 AEG patients in Shanxi province. IVL was selected for further investigation of mutation sites. Esophagus cancer data from TCGA were divided into three groups based on IVL mutations. Differentially-expressed genes were screened and proceeded via Gene Ontology(GO) annotation and Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment analysis. Cell proliferation and migration assays were performed to evaluate the effect of over-expression of IVL on KYSE 150 cells. Mutant IVL oligopeptides were synthesized and used to evaluate the binding activity with TGase1. ResultsFive IVL mutation sites were identified in 22 AEG. GO annotation indicated that these mutations were related to the epithelial differentiation and activity of cell differentiation regulation. KEGG analysis demonstrates that IVL mutations are associated with cAMP signaling pathway. IVL over-expression inhibits the proliferation and migration of esophageal cancer cells(P<0.05). In addition, Q439 H mutant IVL oligopeptides shows lower binding affinity to TGase1 than wild-type oligopeptides.Conclusions The seventh amino acid mutation in IVL repeat motif does weaken IVL binding to TGase1. This mutation may affect tumor-related biological processes and signaling pathways.更多还原