Li Xing, Pan Wei, Qiu Feng, Qiu Zong Yin
Chong Qing University of Medical Sciences, Department of Laboratory Science, Chongqing 400016.
Fen Zi Xi Bao Sheng Wu Xue Bao. 2006 Oct;39(5):399-406.
The subcellular proteome strategy can complement the separation power of two-dimensional electrophoresis, a step for subcellular fractionation is introduced before electrophoresis is conducted, and more proteins will be displayed in two-dimensional gels. A comparative analysis of proteomic profiling of mitochondria and nuclei was conducted between hepatoma cell and hepatocyte, in order to find more information involved in cancer development. The cultured hepatoma cell line QGY-7703 and hepatocyte line LO2 were used as research models in this work. Subcellular fractionation for mitochondrion and nucleus were done by ultracentrifugation, then two-dimensional electrophoresis was applied to the separation of mitochondrial and nuclear proteins, imaging analysis for the selection of differentially-expressed protein spots, and MALDI-TOF-MS for the identification of proteins. 54 spots from electrophoresis gels were selected as differentially-expressed protein for MS analysis which resulted in identification for 22 proteins. Among the 22 differentially-expressed proteins, 17 show a up-regulated expression and 5 show a down-regulated expression. These differentially-expressed proteins found in this work have a wide coverage of functions which are related to energy metabolism, cytosheleton, protein biosynthesis, pre-mRNA splicing and processing, apoptosis regulation. These results imply that cancer cell has experienced a fundamental change in structure and metabolism pattern.
亚细胞蛋白质组策略可以补充二维电泳的分离能力,即在进行电泳之前引入亚细胞分级分离步骤,这样更多的蛋白质将在二维凝胶中显示出来。对肝癌细胞和肝细胞的线粒体和细胞核蛋白质组图谱进行了比较分析,以发现更多与癌症发生发展相关的信息。在这项工作中,使用培养的肝癌细胞系QGY-7703和肝细胞系LO2作为研究模型。通过超速离心对线粒体和细胞核进行亚细胞分级分离,然后应用二维电泳分离线粒体和细胞核蛋白质,通过成像分析选择差异表达的蛋白质斑点,并使用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)鉴定蛋白质。从电泳凝胶中选择54个斑点作为差异表达蛋白质进行质谱分析,结果鉴定出22种蛋白质。在这22种差异表达蛋白质中,17种呈上调表达,5种呈下调表达。在这项工作中发现的这些差异表达蛋白质具有广泛的功能,涉及能量代谢、细胞骨架、蛋白质生物合成、前体mRNA剪接和加工、细胞凋亡调控。这些结果表明癌细胞在结构和代谢模式上经历了根本性的变化。