鼻咽癌组织与正常鼻咽上皮组织的差异磷酸化蛋白质组分析

目的：比较鼻咽癌(nasopharyngeal carcinoma, NPC)组织与正常鼻咽上皮组织磷酸化蛋白质组差异，筛选差异磷酸化蛋白质，为揭示NPC的发病机制提供依据。方法：采用双向凝胶电泳技术(2-DE)分离NPC组织与正常鼻咽上皮组织的总蛋白质，蛋白质转膜后与抗酪氨酸磷酸化抗体进行Western印迹分析，图像分析识别差异磷酸化蛋白质点，电喷雾-四极杆-串联质谱（ESI-Q-TOF MS/MS）鉴定差异的酪氨酸磷酸化蛋白质，采用NetPhos软件预测蛋白质的酪氨酸磷酸化位点，并采用生物信息学方法对差异磷酸化蛋白质的功能和亚细胞定位进行分析。采用Western印迹检测差异蛋白质（phosphatidylethanolamine- binding protein 1）在正常鼻咽黏膜组织和鼻咽癌组织的总蛋白质中的磷酸化水平。结果：建立了NPC组织与正常鼻咽上皮组织的酪氨酸磷酸化蛋白质表达谱，识别了25个差异酪氨酸磷酸化蛋白质，共鉴定了13个差异酪氨酸磷酸化蛋白质，其中7个蛋白质的酪氨酸磷酸化水平在NPC组织中增强，而6个蛋白质的酪氨酸磷酸化水平降低。NetPhos软件预测和生物信息学分析结果显示，13个差异蛋白质均存在酪氨酸磷酸化位点，其功能涉及物质代谢、细胞结构、细胞防御以及信号转导。与正常鼻咽黏膜组织相比较，差异蛋白phosphatidylethanolamine-binding protein 1在鼻咽癌组织中磷酸化表达水平下调。结论： 鉴定了13个可能与NPC发病相关的酪氨酸磷酸化蛋白质，为揭示NPC发病机制提供了科学论依据。

Differential phosphoproteome analysis of nasopharyngeal carcinomaand normal nasopharyngeal epithelial tissues

Objective To compare the difference of phosphoproteome between nasopharyngeal carcinoma(NPC) and normal nasopharyngeal epithelial tissues(NNET) to screening for differential phosphorylated proteins,and provide experimental evidence for elucidating the molecular mechanisms of NPC.Methods Two-dimensional gel electrophoresis(2-DE) coupled with anti-phosphorylated-tyrosine antibody immunoblotting and electrospray ionization-quadrupole time-of-flight MS/MS(ESI-Q-TOF MS/MS) were performed to identify differential tyrosine-phosphorylated proteins in NPC and NNET.NetPhos software was used to predict the tyrosin-phosphorylation sites of identified proteins,and bioinformatics was used to analyze the function and sub-cellular location of identified proteins.A representative result of Western blotting showing changes in the phosphorylation expression levels of phosphatidylethanolamine-binding protein 1 in normal nasopharyngeal mucosal tissues and NPC.Results Phosphoprotein profiles of NPC and NNET were established.Twenty-five differential tyrosine-phosphorylated proteins between the two types of tissues were found,and 13 of them were identified by ESI-Q-TOF MS/MS. Among the identified proteins,tyrosine-phosphorylated levels of 7 proteins were increased,while those of 6 proteins were decreased in NPC.NetPhos software prediction showed that all of 13 proteins contain the tyrosine phosphorylation sites.Compared with normal nasopharyngeal mucosal tissues,the phosphorylation expression levels of phosphatidylethanolamine-binding protein 1 in nasopharyngeal carcinoma are decreased.Conclusion Thirteen differential tyrosine-phosphorylated proteins possibly involved in NPC carcinogenesis were identified,and the data will be helpful to elucidate the molecular mechanisms of NPC.