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Abstract：
OBJECTIVE: This study aims to elucidate the functional role of IQGAP1 phosphorylation modification mediated by the SOX4/MAPK1 regulatory axis in developing pancreatic cancer through phosphoproteomics analysis.
METHODS: Proteomics and phosphoproteomics data of pancreatic cancer were obtained from the Clinical Proteomic Tumor Analysis Consortium (CPTAC) database. Differential analysis, kinase-substrate enrichment analysis (KSEA), and independent prognosis analysis were performed on these datasets. Subtype analysis of pancreatic cancer patients was conducted based on the expression of prognostic-related proteins, and the prognosis of different subtypes was evaluated through prognosis analysis. Differential analysis of proteins in different subtypes was performed to identify differential proteins in the high-risk subtype. Clinical correlation analysis was conducted based on the expression of prognostic-related proteins, pancreatic cancer typing results, and clinical characteristics in the pancreatic cancer proteomics dataset. Functional pathway enrichment analysis was performed using GSEA/GO/KEGG, and most module proteins correlated with pancreatic cancer were selected using WGCNA analysis. In cell experiments, pancreatic cancer cells were grouped, and the expression levels of SOX4, MAPK1, and the phosphorylation level of IQGAP1 were detected by RT-qPCR and Western blot experiments. The effect of SOX4 on MAPK1 promoter transcriptional activity was assessed using a dual-luciferase assay, and the enrichment of SOX4 on the MAPK1 promoter was examined using a ChIP assay. The proliferation, migration, and invasion functions of grouped pancreatic cancer cells were assessed using CCK-8, colony formation, and Transwell assays. In animal experiments, the impact of SOX4 on tumor growth and metastasis through the regulation of MAPK1-IQGAP1 phosphorylation modification was studied by constructing subcutaneous and orthotopic pancreatic cancer xenograft models, as well as a liver metastasis model in nude mice.
RESULTS: Phosphoproteomics and proteomics data analysis revealed that the kinase MAPK1 may play an important role in pancreatic cancer progression by promoting IQGAP1 phosphorylation modification. Proteomics analysis classified pancreatic cancer patients into two subtypes, C1 and C2, where the high-risk C2 subtype was associated with poor prognosis, malignant tumor typing, and enriched tumor-related pathways. SOX4 may promote the occurrence of the high-risk C2 subtype of pancreatic cancer by regulating MAPK1-IQGAP1 phosphorylation modification. In vitro cell experiments confirmed that SOX4 promoted IQGAP1 phosphorylation modification by activating MAPK1 transcription while silencing SOX4 inhibited the proliferation, migration, and invasion of pancreatic cancer cells by reducing the phosphorylation level of MAPK1-IQGAP1. In vivo, animal experiments further confirmed that silencing SOX4 suppressed the growth and metastasis of pancreatic cancer by reducing the phosphorylation level of MAPK1-IQGAP1.
CONCLUSIONS: The findings of this study suggest that SOX4 promotes the phosphorylation modification of IQGAP1 by activating MAPK1 transcription, thereby facilitating the growth and metastasis of pancreatic cancer.
METHODS: Proteomics and phosphoproteomics data of pancreatic cancer were obtained from the Clinical Proteomic Tumor Analysis Consortium (CPTAC) database. Differential analysis, kinase-substrate enrichment analysis (KSEA), and independent prognosis analysis were performed on these datasets. Subtype analysis of pancreatic cancer patients was conducted based on the expression of prognostic-related proteins, and the prognosis of different subtypes was evaluated through prognosis analysis. Differential analysis of proteins in different subtypes was performed to identify differential proteins in the high-risk subtype. Clinical correlation analysis was conducted based on the expression of prognostic-related proteins, pancreatic cancer typing results, and clinical characteristics in the pancreatic cancer proteomics dataset. Functional pathway enrichment analysis was performed using GSEA/GO/KEGG, and most module proteins correlated with pancreatic cancer were selected using WGCNA analysis. In cell experiments, pancreatic cancer cells were grouped, and the expression levels of SOX4, MAPK1, and the phosphorylation level of IQGAP1 were detected by RT-qPCR and Western blot experiments. The effect of SOX4 on MAPK1 promoter transcriptional activity was assessed using a dual-luciferase assay, and the enrichment of SOX4 on the MAPK1 promoter was examined using a ChIP assay. The proliferation, migration, and invasion functions of grouped pancreatic cancer cells were assessed using CCK-8, colony formation, and Transwell assays. In animal experiments, the impact of SOX4 on tumor growth and metastasis through the regulation of MAPK1-IQGAP1 phosphorylation modification was studied by constructing subcutaneous and orthotopic pancreatic cancer xenograft models, as well as a liver metastasis model in nude mice.
RESULTS: Phosphoproteomics and proteomics data analysis revealed that the kinase MAPK1 may play an important role in pancreatic cancer progression by promoting IQGAP1 phosphorylation modification. Proteomics analysis classified pancreatic cancer patients into two subtypes, C1 and C2, where the high-risk C2 subtype was associated with poor prognosis, malignant tumor typing, and enriched tumor-related pathways. SOX4 may promote the occurrence of the high-risk C2 subtype of pancreatic cancer by regulating MAPK1-IQGAP1 phosphorylation modification. In vitro cell experiments confirmed that SOX4 promoted IQGAP1 phosphorylation modification by activating MAPK1 transcription while silencing SOX4 inhibited the proliferation, migration, and invasion of pancreatic cancer cells by reducing the phosphorylation level of MAPK1-IQGAP1. In vivo, animal experiments further confirmed that silencing SOX4 suppressed the growth and metastasis of pancreatic cancer by reducing the phosphorylation level of MAPK1-IQGAP1.
CONCLUSIONS: The findings of this study suggest that SOX4 promotes the phosphorylation modification of IQGAP1 by activating MAPK1 transcription, thereby facilitating the growth and metastasis of pancreatic cancer.
摘要：
目的：本研究旨在通过磷酸化蛋白质组学分析阐明SOX4/MAPK1调节轴介导的IQGAP1磷酸化修饰在胰腺癌发展中的功能作用。
方法：胰腺癌的蛋白质组学和磷酸蛋白质组学数据来自临床蛋白质组学肿瘤分析联盟（CPTAC）数据库。差异分析,激酶-底物富集分析(KSEA)，并对这些数据集进行独立预后分析.根据预后相关蛋白的表达对胰腺癌患者进行亚型分析,并通过预后分析评估不同亚型的预后。对不同亚型中的蛋白质进行差异分析以鉴定高风险亚型中的差异蛋白质。根据预后相关蛋白的表达进行临床相关性分析,胰腺癌分型结果,和胰腺癌蛋白质组学数据集中的临床特征。使用GSEA/GO/KEGG进行功能途径富集分析，并且使用WGCNA分析选择与胰腺癌相关的大多数模块蛋白。在细胞实验中,胰腺癌细胞被分组，通过RT-qPCR和Westernblot实验检测SOX4，MAPK1的表达水平和IQGAP1的磷酸化水平。SOX4对MAPK1启动子转录活性的影响使用双荧光素酶测定来评估，并使用ChIP测定法检查MAPK1启动子上SOX4的富集。扩散,迁移,使用CCK-8，集落形成评估分组胰腺癌细胞的侵袭功能，和Transwell分析。在动物实验中,通过构建皮下和原位胰腺癌异种移植模型，研究了SOX4通过调节MAPK1-IQGAP1磷酸化修饰对肿瘤生长和转移的影响，以及裸鼠肝转移模型。
结果：磷酸化蛋白质组学和蛋白质组学数据分析显示，激酶MAPK1可能通过促进IQGAP1磷酸化修饰在胰腺癌进展中发挥重要作用。蛋白质组学分析将胰腺癌患者分为两种亚型，C1和C2，其中高风险C2亚型与不良预后相关，恶性肿瘤分型,和丰富的肿瘤相关通路。SOX4可能通过调节MAPK1-IQGAP1磷酸化修饰促进胰腺癌高危C2亚型的发生。体外细胞实验证实，SOX4通过激活MAPK1转录促进IQGAP1磷酸化修饰，沉默SOX4抑制增殖，迁移,通过降低MAPK1-IQGAP1的磷酸化水平来实现胰腺癌细胞的侵袭。在体内,动物实验进一步证实沉默SOX4通过降低MAPK1-IQGAP1的磷酸化水平抑制胰腺癌的生长和转移。
结论：这项研究的结果表明，SOX4通过激活MAPK1转录促进IQGAP1的磷酸化修饰，从而促进胰腺癌的生长和转移。
方法：胰腺癌的蛋白质组学和磷酸蛋白质组学数据来自临床蛋白质组学肿瘤分析联盟（CPTAC）数据库。差异分析,激酶-底物富集分析(KSEA)，并对这些数据集进行独立预后分析.根据预后相关蛋白的表达对胰腺癌患者进行亚型分析,并通过预后分析评估不同亚型的预后。对不同亚型中的蛋白质进行差异分析以鉴定高风险亚型中的差异蛋白质。根据预后相关蛋白的表达进行临床相关性分析,胰腺癌分型结果,和胰腺癌蛋白质组学数据集中的临床特征。使用GSEA/GO/KEGG进行功能途径富集分析，并且使用WGCNA分析选择与胰腺癌相关的大多数模块蛋白。在细胞实验中,胰腺癌细胞被分组，通过RT-qPCR和Westernblot实验检测SOX4，MAPK1的表达水平和IQGAP1的磷酸化水平。SOX4对MAPK1启动子转录活性的影响使用双荧光素酶测定来评估，并使用ChIP测定法检查MAPK1启动子上SOX4的富集。扩散,迁移,使用CCK-8，集落形成评估分组胰腺癌细胞的侵袭功能，和Transwell分析。在动物实验中,通过构建皮下和原位胰腺癌异种移植模型，研究了SOX4通过调节MAPK1-IQGAP1磷酸化修饰对肿瘤生长和转移的影响，以及裸鼠肝转移模型。
结果：磷酸化蛋白质组学和蛋白质组学数据分析显示，激酶MAPK1可能通过促进IQGAP1磷酸化修饰在胰腺癌进展中发挥重要作用。蛋白质组学分析将胰腺癌患者分为两种亚型，C1和C2，其中高风险C2亚型与不良预后相关，恶性肿瘤分型,和丰富的肿瘤相关通路。SOX4可能通过调节MAPK1-IQGAP1磷酸化修饰促进胰腺癌高危C2亚型的发生。体外细胞实验证实，SOX4通过激活MAPK1转录促进IQGAP1磷酸化修饰，沉默SOX4抑制增殖，迁移,通过降低MAPK1-IQGAP1的磷酸化水平来实现胰腺癌细胞的侵袭。在体内,动物实验进一步证实沉默SOX4通过降低MAPK1-IQGAP1的磷酸化水平抑制胰腺癌的生长和转移。
结论：这项研究的结果表明，SOX4通过激活MAPK1转录促进IQGAP1的磷酸化修饰，从而促进胰腺癌的生长和转移。
