Zanivan Sara, Gnad Florian, Wickström Sara A, Geiger Tami, Macek Boris, Cox Jürgen, Fässler Reinhard, Mann Matthias
Department of Proteomics and Signal Transduction, Max-Planck-Institute for Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany.
J Proteome Res. 2008 Dec;7(12):5314-26. doi: 10.1021/pr800599n.
Kinases play a prominent role in tumor development, pointing to the presence of specific phosphorylation patterns in tumor tissues. Here, we investigate whether recently developed high resolution mass spectrometric (MS) methods for proteome and phosphoproteome analysis can also be applied to solid tumors. As tumor model, we used TG3 mutant mice carrying skin melanomas. At total of 100 microg of solid tumor lysate yielded a melanoma proteome of 4443 identified proteins, including at least 88 putative melanoma markers previously found by cDNA microarray technology. Analysis of 2 mg of lysate from dissected melanoma with titansphere chromatography and 8 mg with strong cation exchange together resulted in the identification of more than 5600 phosphorylation sites on 2250 proteins. The phosphoproteome included many hits from pathways important in melanoma. One-month storage at -80 degrees C did not significantly decrease the number of identified phosphorylation sites. Thus, solid tumor can be analyzed by MS-based proteomics with similar efficiency as cell culture models and in amounts compatible with biopsies.
激酶在肿瘤发展过程中发挥着重要作用，这表明肿瘤组织中存在特定的磷酸化模式。在此，我们研究最近开发的用于蛋白质组和磷酸化蛋白质组分析的高分辨率质谱（MS）方法是否也可应用于实体肿瘤。作为肿瘤模型，我们使用了携带皮肤黑色素瘤的TG3突变小鼠。100微克实体肿瘤裂解物共产生了一个包含4443种已鉴定蛋白质的黑色素瘤蛋白质组，其中包括至少88种先前通过cDNA微阵列技术发现的假定黑色素瘤标志物。用钛球色谱法分析2毫克解剖后的黑色素瘤裂解物，并用强阳离子交换法分析8毫克裂解物，共鉴定出2250种蛋白质上的5600多个磷酸化位点。磷酸化蛋白质组包含许多来自黑色素瘤重要通路的命中结果。在-80℃下储存一个月并未显著减少已鉴定磷酸化位点的数量。因此，基于质谱的蛋白质组学可以以与细胞培养模型相似的效率分析实体肿瘤，并且分析量与活检兼容。
