对于有丝分裂和减数分裂而言,着丝粒是不可或缺的重要功能元件。近几年的研究表明,着丝粒蛋白F(Centromere Protein F,CENPF)是细胞周期调控的关键蛋白。CENPF是一种分子量为367kDa的核定位蛋白,它在有丝分裂前期开始增加,在有丝分裂期...对于有丝分裂和减数分裂而言,着丝粒是不可或缺的重要功能元件。近几年的研究表明,着丝粒蛋白F(Centromere Protein F,CENPF)是细胞周期调控的关键蛋白。CENPF是一种分子量为367kDa的核定位蛋白,它在有丝分裂前期开始增加,在有丝分裂期定位于动粒,末期开始迅速降解。CENPF缺失会造成有丝分裂缺陷,包括纺锤体无法正常组装、染色体非正常排列和分离,甚至细胞死亡。同时,CENPF可能与纺锤体组装检验点(Spindle Assembly Checkpoint,SAC)的活性有关。CENPF功能的发挥取决于法尼基化修饰,CENPF的过表达则可能导致肿瘤的发生,通过法尼基转移酶抑制剂(Farnesyltransferase Inhibitor,FTI)处理细胞则会抑制肿瘤细胞的增殖。同时,CENPF对小鼠早期胚胎的发育也具有重要的调控作用。对近些年CENPF的研究进展进行了综述,并提出了尚待解决的关键问题,希望可以为后续的研究提供一些思路。展开更多
Background:N6-methyladenosine(m^(6)A)modification is the most common modification that occurs in eukaryotes.Although substantial effort has been made in the prevention and treatment of gastric cancer(GC)in recent year...Background:N6-methyladenosine(m^(6)A)modification is the most common modification that occurs in eukaryotes.Although substantial effort has been made in the prevention and treatment of gastric cancer(GC)in recent years,the prognosis of GC patients remains unsatisfactory.The regulatory mechanism between m^(6)A modification and GC development needs to be elucidated.In this study,we examined m^(6)A modification and the downstream mechanism in GC.Methods:Dot blotting assays,The Cancer Genome Atlas analysis,and quantitative real‑time PCR(qRT-PCR)were used to measure the m^(6)A levels in GC tissues.Methylated RNA-immunoprecipitation sequencing and RNA sequencingwere performed to identify the targets ofm^(6)Amodification.Western blotting,Transwell,wound healing,and angiogenesis assays were conducted to examine the role of centromere protein F(CENPF)in GC in vitro.Xenograft,immunohistochemistry,and in vivo metastasis experiments were conducted to examine the role of CENPF in GC in vivo.Methylated RNA-immunoprecipitation-qPCR,RNA immunoprecipitation-qPCR and RNA pulldown assays were used to verify the m^(6)A modification sites of CENPF.Gain/loss-of-function and rescue experiments were conducted to determine the relationship between CENPF and the mitogen-activated protein kinase(MAPK)signaling pathway in GC cells.Coimmunoprecipitation,mass spectrometry,qRT-PCR,and immunofluorescence assays were performed to explore the proteins that interact with CENPF and elucidate the regulatory mechanisms between them.Results:CENPF was upregulated in GC and facilitated the metastasis of GC both in vitro and in vivo.Mechanistically,increasedm^(6)A modification of CENPF was mediated by methyltransferase 3,and this modified molecule could be recognized by heterogeneous nuclear ribonucleoprotein A2/B1(HNRNPA2B1),thereby promoting its mRNA stability.In addition,the metastatic phenotype of CENPF was dependent on the MAPK signaling pathway.Furthermore,CENPF could bind to FAK and promote its localization in the cytoplasm.Moreover,we discovered that high expression of CENPF was related to lymphatic invasion and overall survival in GC patients.Conclusions:Our findings revealed that increased m^(6)A modification of CENPF facilitates the metastasis and angiogenesis of GC through the CENPF/FAK/MAPK and epithelial-mesenchymal transition axis.CENPF expression was correlated with the clinical features of GC patients;therefore,CENPF may serve as a prognostic marker of GC.展开更多
目的探究着丝粒蛋白F(centromere protein F,CENPF)在胰腺导管腺癌(pancreatic ductual adenocarcinoma,PDAC)中的表达及其与临床病理特征和预后的关系。方法基于美国国家生物信息中心(National Center for Biotechnology Information,N...目的探究着丝粒蛋白F(centromere protein F,CENPF)在胰腺导管腺癌(pancreatic ductual adenocarcinoma,PDAC)中的表达及其与临床病理特征和预后的关系。方法基于美国国家生物信息中心(National Center for Biotechnology Information,NCBI)的微阵列/基因谱公共数据库(Gene Expression Omnibus,GEO),运用GEO2R、维恩、Cytoscape及GEPIA软件筛选出PDAC中可疑差异基因CENPF;基于癌症基因组图谱(The Cancer Genome Atlas,TCGA)及基因组织表达数据库(The Genotype-Tissue Expression,GTEx),通过NCBI网络、GEPIA、Ualcan、Oncomine、TIMER软件及Kaplan-Meier在线生存分析工具,从信使核糖核酸(messenger RNA)层面分析其表达与免疫细胞浸润的关系及其可能的分子机制。同时收集2007~2021年间经福建医科大学附属第一医院病理科确诊的胰腺导管腺癌手术标本121例,从组织蛋白层面分析CENPF与PDAC临床病理特征及预后的关系。结果CENPF基因在人类多种癌症中表达异常,在PDAC中,癌组织中的表达水平显著高于癌旁正常胰腺组织(P<0.05),并且与PDAC组织学分级(P<0.05)、预后(P=0.0038)有关。免疫组化显示CENPF的表达与PDAC的神经侵犯(P=0.036)、TNM分期(P=0.041)、淋巴结转移(P=0.023)、分化程度(P=0.020)、总生存期有关(Log-rank=18.608,P=0.000016),CENPF高表达(HR=2.654,95%CI=1.373~5.131,P=0.004)是PDAC患者预后差的独立危险因素。CENPF联合其他关键模块基因在PDAC中,主要富集于外泌体、细胞外基质等,并与丝氨酸内肽酶活性、金属肽链内切酶活性有关等。CENPF在胰腺导管腺癌中的作用通路主要与细胞周期、P53通路、泛素介导的蛋白水解等相关,并与P53、MDM2在PDAC中联合发挥作用。免疫浸润研究表明CENPF表达与CD4+T淋巴细胞浸润负相关、与树突细胞的浸润正相关(均P<0.05)。结论CENPF可能参与PDAC疾病的进程,其高表达与PDAC预后不良相关,该研究有望为胰腺导管腺癌的防治提供更多科学基础。展开更多
基金Special Foundation for National Science and Technology Basic Research Program of China,Grant/Award Number:2019FY101104National Natural Science Foundation of China,Grant/Award Numbers:81871946,82072708+6 种基金Primary Research&Development Plan of Jiangsu Province,Grant/Award Number:BE2016786Program for Development of Innovative Research Team in the First Affiliated Hospital of NJMUPriority Academic Program Development of Jiangsu Higher Education Institutions,Grant/Award Number:JX10231801Jiangsu Key Medical Discipline,Grant/Award Number:ZDXKA2016005Jiangsu Key Lab of Cancer Biomarkers,Prevention and TreatmentCollaborative Innovation Centre for Cancer Personalized MedicineNanjing Medical University。
文摘Background:N6-methyladenosine(m^(6)A)modification is the most common modification that occurs in eukaryotes.Although substantial effort has been made in the prevention and treatment of gastric cancer(GC)in recent years,the prognosis of GC patients remains unsatisfactory.The regulatory mechanism between m^(6)A modification and GC development needs to be elucidated.In this study,we examined m^(6)A modification and the downstream mechanism in GC.Methods:Dot blotting assays,The Cancer Genome Atlas analysis,and quantitative real‑time PCR(qRT-PCR)were used to measure the m^(6)A levels in GC tissues.Methylated RNA-immunoprecipitation sequencing and RNA sequencingwere performed to identify the targets ofm^(6)Amodification.Western blotting,Transwell,wound healing,and angiogenesis assays were conducted to examine the role of centromere protein F(CENPF)in GC in vitro.Xenograft,immunohistochemistry,and in vivo metastasis experiments were conducted to examine the role of CENPF in GC in vivo.Methylated RNA-immunoprecipitation-qPCR,RNA immunoprecipitation-qPCR and RNA pulldown assays were used to verify the m^(6)A modification sites of CENPF.Gain/loss-of-function and rescue experiments were conducted to determine the relationship between CENPF and the mitogen-activated protein kinase(MAPK)signaling pathway in GC cells.Coimmunoprecipitation,mass spectrometry,qRT-PCR,and immunofluorescence assays were performed to explore the proteins that interact with CENPF and elucidate the regulatory mechanisms between them.Results:CENPF was upregulated in GC and facilitated the metastasis of GC both in vitro and in vivo.Mechanistically,increasedm^(6)A modification of CENPF was mediated by methyltransferase 3,and this modified molecule could be recognized by heterogeneous nuclear ribonucleoprotein A2/B1(HNRNPA2B1),thereby promoting its mRNA stability.In addition,the metastatic phenotype of CENPF was dependent on the MAPK signaling pathway.Furthermore,CENPF could bind to FAK and promote its localization in the cytoplasm.Moreover,we discovered that high expression of CENPF was related to lymphatic invasion and overall survival in GC patients.Conclusions:Our findings revealed that increased m^(6)A modification of CENPF facilitates the metastasis and angiogenesis of GC through the CENPF/FAK/MAPK and epithelial-mesenchymal transition axis.CENPF expression was correlated with the clinical features of GC patients;therefore,CENPF may serve as a prognostic marker of GC.
文摘目的探究着丝粒蛋白F(centromere protein F,CENPF)在胰腺导管腺癌(pancreatic ductual adenocarcinoma,PDAC)中的表达及其与临床病理特征和预后的关系。方法基于美国国家生物信息中心(National Center for Biotechnology Information,NCBI)的微阵列/基因谱公共数据库(Gene Expression Omnibus,GEO),运用GEO2R、维恩、Cytoscape及GEPIA软件筛选出PDAC中可疑差异基因CENPF;基于癌症基因组图谱(The Cancer Genome Atlas,TCGA)及基因组织表达数据库(The Genotype-Tissue Expression,GTEx),通过NCBI网络、GEPIA、Ualcan、Oncomine、TIMER软件及Kaplan-Meier在线生存分析工具,从信使核糖核酸(messenger RNA)层面分析其表达与免疫细胞浸润的关系及其可能的分子机制。同时收集2007~2021年间经福建医科大学附属第一医院病理科确诊的胰腺导管腺癌手术标本121例,从组织蛋白层面分析CENPF与PDAC临床病理特征及预后的关系。结果CENPF基因在人类多种癌症中表达异常,在PDAC中,癌组织中的表达水平显著高于癌旁正常胰腺组织(P<0.05),并且与PDAC组织学分级(P<0.05)、预后(P=0.0038)有关。免疫组化显示CENPF的表达与PDAC的神经侵犯(P=0.036)、TNM分期(P=0.041)、淋巴结转移(P=0.023)、分化程度(P=0.020)、总生存期有关(Log-rank=18.608,P=0.000016),CENPF高表达(HR=2.654,95%CI=1.373~5.131,P=0.004)是PDAC患者预后差的独立危险因素。CENPF联合其他关键模块基因在PDAC中,主要富集于外泌体、细胞外基质等,并与丝氨酸内肽酶活性、金属肽链内切酶活性有关等。CENPF在胰腺导管腺癌中的作用通路主要与细胞周期、P53通路、泛素介导的蛋白水解等相关,并与P53、MDM2在PDAC中联合发挥作用。免疫浸润研究表明CENPF表达与CD4+T淋巴细胞浸润负相关、与树突细胞的浸润正相关(均P<0.05)。结论CENPF可能参与PDAC疾病的进程,其高表达与PDAC预后不良相关,该研究有望为胰腺导管腺癌的防治提供更多科学基础。
