蛋白质的赖氨酸甲基化是一种在细胞内部起着关键调控作用的翻译后修饰类型。对不同生理状态下赖氨酸甲基化的位点占有率进行测定分析，对于解析赖氨酸甲基化对生理过程的动态调控非常重要，但到目前为止，还没有相应的技术手段对赖氨酸甲基化的位点占有率进行高通量测定分析。针对这一现状，本项目拟发展一种综合了hM-SILAC标记、二甲基化衍生和PRM分析的赖氨酸甲基化位点占有率的测定新方法，实现位点占有率层次上的精准测定分析；拟将发展的技术手段用于肿瘤细胞耐药过程中赖氨酸甲基化位点占有率的差异分析，探索赖氨酸甲基化的位点占有率与肿瘤细胞耐药性之间的关联。本项目的开展实施将有效地提升我国在赖氨酸甲基化定量分析方面的技术水平。

Lysine methylation is an important post-translational-modification (PTM) that plays crucial roles in diverse physiological and pathological processes. Direct measurement of the methylation stoichiometry can unambiguously reveal whether the signal-induced alteration is regulated by methyltransferase/demethylase activity or by transcriptional regulation to modify protein abundance, however, no methodology is available for the high-throughput stoichiometry analysis of lysine methylation. Thus, it is of great urgency to overcome the current deficiency in accessing the stoichiometry of lysine methylation. To this end, we propose to develop an integrated approach that utilizes hM-SILAC and dimethyl labeling to determine the absolute stoichiometry of lysine methylation on a large scale. Furthermore, the newly developed methods will be utilized to the analysis of protein methylation in drug-resistance cancer cells, which could reveal the connections between lysine methylation and drug-resistance. Based on the above results, we will further investigate the relationship among methylated proteins and drug-resistance. The outcome of this project will greatly improve the technology advancement in quantitative analysis of lysine methylation.