转移潜能不同的人前列腺癌细胞系差异膜蛋白质组学的分析

Differential plasma membrane proteomic analysis of human prostate cancer cell line with various metastasis potency

目的:通过应用鸟枪法蛋白质组学技术发现早期诊断和逆转前列腺癌转移的分子标志。方法:应用鸟枪法蛋白质组学技术,比较人高转移前列腺癌细胞株(PC-3M)和人低转移前列腺癌细胞株(PC-3)间膜蛋白表达的差异。用SDS-PAGE胶分离膜蛋白,并且进行胰酶消化,用反相液相色谱分离肽混合物后进行ESI-MS/MS鉴定,用数据库查询结合生物信息学技术比较分析并识别细胞间的差异表达膜蛋白。结果:1)利用Gel-1DLC-MS/MS分析,在严格的过滤参数条件下鉴定了PC-3和PC-3M两种细胞中的蛋白,在PC-3细胞中鉴定了2172个蛋白,在PC-3M细胞中鉴定了2023个蛋白。2)用GOA分析软件进行分类,PC-3细胞中1499个蛋白为已知细胞组分,其中564(38.13%)个蛋白为膜蛋白或者膜相关蛋白;PC-3M细胞中1391个蛋白为已知细胞组分,细胞组分中527(38.30%)个为膜蛋白或者膜相关蛋白。几个假设蛋白和cDNA序列也被预测。3)用蛋白名称和IPI数据库号对比分析后发现有161个蛋白质仅在PC-3前列腺癌细胞株中检测到有表达,135个蛋白质仅在PC-3M中检测到有表达。结论:人高、低转移前列腺癌细胞株的膜蛋白表达存在明显差异,为进一步发现前列腺癌转移相关分子和阐明前列腺癌侵袭转移的分子机制提供实验证据。

OBJECTIVE： To explore the mechanism of prostate cancer metastasis and search for the potential markers for early diagnosing and reversing prostate cancer metastasis by shotgun-MS. METHODS： By using the shotgun strategy, the differential plasma membrane proteomic analysis was conducted in two human prostate cancer cell lines which were high metastasis potentials PC-3M cells and low metastasis potentials PC-3 cells. The plasma membrane （PM） proteins were separated by SDS-PAGE and digested with trypsin, separation of peptide mixture by RP-HPLC followed by ESI-MS/MS identification. The differentially expressed proteins of the two cell lines were analyzed by the data base query and bioirformatics technique. RESISTS.. The proteins of the two cell lines were separated by Gel- 1DLC- MS/ MS. The proteins were identified under a stringent filter condition in both PC-3 and PC-3M cells pools, in which 2 172 and 2 023 entire proteins were found, respectively. A total of 2 172 proteins in PC-3 cells were identified, of which 1 499 had a gene ontology （GO） annotation indicating a cellular component, and 564（38. 13%） of the latter were known membrane proteins or membrane-associated proteins. Among the 2 023 identified proteins of PC-3M cells, 1 391 had a GO annotation indicating a cellular component, and 527（38.30%） of the latter were known membrane proteins or membrane-associated proteins. In addition to the proteins known to be associated with the membrane, several hypothetical proteins and cDNA sequences were also identified. The protein name and IPI data analysis of human re- vealed that 217 protein spots had significant differential expressions in PC-3 and PC-3M, 161 protein spots were only detected in PC-3 cells, and 135 protein spots were only detected in PC-3M cells. CONCLUSlONS：It is suggested that an obviously differential membrane proteomic expression exists between the human highand low-metastatic prostate cancer cell lines. It will be helpful to further understand the molecular mechanisms of prostate cancer invasion and metastasis, and provide new experimental evidence for searching metastatic-related molecule of prostate cancer.