A novel metastatic promoter CEMIP and its downstream molecular targets and signaling pathway of cellular migration and invasion in SCLC cells based on proteome analysis
A novel metastatic promoter CEMIP and its downstream molecular targets and signaling pathway of cellular migration and invasion in SCLC cells based on proteome analysis
复制标题
基于蛋白质组分析的SCLC细胞新型转移启动子CEMIP及其下游分子靶点和细胞迁移侵袭信号通路
DOI：
10.1007/s00432-020-03308-5
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发表时间：
2020-07-09
影响因子：
3.6
通讯作者：
Yang, Jie
中科院分区：
文献类型：
--
作者：
Li, Li;Pan, Yingxing;Yang, Jie
目的转移是几乎所有小细胞肺癌(SCLC)患者都会发生的不可避免的事件。然而，分子驱动因素还没有被阐明。最近，一种名为细胞迁移诱导透明质酸酶(CEMIP)的新的水解酶在许多肿瘤中同时触发了迁移和侵袭，但不是小细胞肺癌。因此，在本研究中，我们验证了CEMIP促进小细胞肺癌的迁移和侵袭，并应用蛋白质组学分析筛选出与CEMIP调控相关的潜在靶点和信号通路。方法采用免疫荧光方法检测CEMIP在小细胞肺癌及其配对的正常组织中的表达。RT-qPCR和Western印迹(WB)检测细胞CEMIP和EMT标志物的蛋白和mRNA表达。采用慢病毒CEMIP-shRNAs和CEMIP表达载体进行表达调控。通过Transwell法检测细胞迁移和侵袭能力的变化。结果小细胞肺癌组织中CEMIP的表达水平较正常组织高1.64 ± 0.16倍。降低CEMIP在小细胞肺癌细胞H446上的表达可降低细胞的迁移和侵袭能力，而通过在H1688上过表达CEMIP来促进细胞的迁移和侵袭。蛋白质组学和生物信息学分析表明，共有215个相对表达增强或降低的差异表达蛋白达到了p值为 ≤ 0.05的1.2倍变化阈值。显著上调的DEPS包括一个未知的多肽序列(由cDNA FLJ52096编码)SPICE1和CryAB，而S100A6的表达则大幅下调。DEPS主要富含细胞成分的小凹、生物学过程中的钙离子结合和上皮细胞的分子功能迁移。结果提示，DEPS主要作用于转化生长因子β、GABA能突触和丝裂原活化蛋白K信号转导通路。为CEMIP诱导细胞迁移和侵袭SCLC提供了可能的分子机制线索和潜在的下游靶点。
PurposeMetastasis is an unavoidable event happened among almost all small cell lung cancer (SCLC) patients. However, the molecular driven factors have not been elucidated. Recently, a novel hydrolase called cell migration inducing hyaluronidase (CEMIP) triggered both migration and invasion in many tumors but not SCLC. Therefore, in this study, we verified that CEMIP promoted migration and invasion in SCLC and applied proteomics analysis to screen out potential target profiles and the signaling pathway related to CEMIP regulation.MethodImmunofluorescence was conducted to exam the expression of CEMIP on SCLC and paired adjacent normal tissues among enrollment. RT-qPCR and Western blot (WB) assays were conducted to valuate cellular protein and mRNA expression of CEMIP and EMT markers. Lentivirus-CEMIP-shRNAs and CEMIP plasmid were used for expression manipulating. Changes of cellular migration and invasion were tested through transwell assays. Tandem Mass Tag (TMT) peptide labeling coupled with LC–MS/MS was used for quantifying proteins affected by reducing expression of CEMIP on H446 cells.ResultsThe expression of CEMIP showed 1.64 ± 0.16-fold higher in SCLC tissues than their normal counterpart. Decreasing the expression of CEMIP on SCLC cells H446 regressed both cellular migration and invasion ability, whereas the promoting cellular migration and invasion was investigated through over-expressing CEMIP on H1688. Proteomic and bioinformatics analysis revealed that total 215 differentially expressed proteins (DEPs) that either their increasing or decreasing relative expression met threshold of 1.2-fold changes with p value ≤ 0.05. The dramatic up-regulated DEPs included an unidentified peptide sequence (encoded by cDNA FLJ52096) SPICE1 and CRYAB, while the expression of S100A6 was largely down-regulated. DEPs mainly enriched on caveolae of cellular component, calcium ion binding of biological process and epithelial cell migration of molecular function. KEGG enrichment indicated that DEPs mainly exerted their function on TGF-β, GABAergic synapse and MAPK signaling pathway.ConclusionIt is the first report illustrating that CEMIP might be one of the metastatic triggers in SCLC. And also, it provided possible molecular mechanism cue and potential downstream target on CEMIP-induced cellular migration and invasion on SCLC.