Comparative proteomic analysis of protein methylation provides insight into the resistance of hepatocellular carcinoma to 5-fluorouracil
Comparative proteomic analysis of protein methylation provides insight into the resistance of hepatocellular carcinoma to 5-fluorouracil
复制标题
蛋白质甲基化的比较蛋白质组学分析可深入了解肝细胞癌对 5-氟尿嘧啶的耐药性
DOI：
10.1016/j.jprot.2020.103738
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发表时间：
2020-05-15
影响因子：
3.3
通讯作者：
Ye, Mingliang
中科院分区：
文献类型：
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作者：
Liu, Zhen;Wang, Qi;Ye, Mingliang
蛋白质甲基化是一种常见的翻译后修饰，参与多种生物学过程，包括信号转导、转录调控、DNA修复、基因激活、基因阻遏和RNA加工。由于技术条件的限制，目前对癌细胞中蛋白质甲基化的研究还不多，这阻碍了我们对蛋白质甲基化在肿瘤耐药中作用的理解。本研究采用SPE-SCX技术对5-氟尿嘧啶（5-Fu）耐药细胞系Bel/5-Fu及其亲本细胞系的甲基化蛋白质组进行了分析。我们在Bel细胞中鉴定了294个位点上的313种甲基化形式，在Bel/5-Fu细胞中鉴定了260个位点上的294种甲基化形式，具有较高的定位置信度。此外，我们对251种甲基化形式进行了定量，发现77种甲基化形式发生了显著变化。在用蛋白质丰度归一化后，确定了89种甲基化形式，其中位点化学计量学发生了显著变化。这些显著改变的甲基化位点的序列特征是不同的。基因本体分析表明，这些显著改变的甲基化蛋白主要参与翻译和转录等生物学过程。总之，我们的研究结果表明，发生在肝细胞癌中的蛋白质甲基化可能在需要耐药方面发挥关键作用。意义：在癌细胞中获得的耐药性已被认为是癌症治疗的主要挑战。由于其复杂性，其分子机制在很大程度上仍然未知。确定关键标志物将提高我们对机制的理解，对于开发克服耐药性的新治疗策略至关重要。目前，越来越多的蛋白质组学和磷酸化蛋白质组学研究被报道用于研究耐药机制。但与耐药性相关的甲基蛋白质组学研究尚未见报道。本研究采用SPE-SCX无标记定量蛋白质组学方法对耐药细胞系Bel/5-Fu和敏感细胞系Bel的甲基化蛋白质组进行了分析。通过定性和定量分析，发现两种细胞系的甲基化位点序列特征存在明显差异。结果提示，某些甲基转移酶可能在耐药性的调控中起重要作用。我们还通过标准化蛋白质丰度进行甲基位点化学计量分析。结果表明，Bel细胞中存在89种甲基化形式，位点化学计量学发生了显著变化，这可能与Bel细胞向抗性细胞的发展有关。我们的甲基化蛋白质组数据集将有助于揭示肝细胞癌获得耐药的新分子机制。
Protein methylation is one of the common post-translational modifications involved in diverse biological processes including signal transduction, transcriptional regulation, DNA repairing, gene activation, gene repression, and RNA processing. Due to technique limitation, the investigation of protein methylation in cancer cells is not well achieved, which hinders our understanding of the contribution of protein methylation to drug resistance. In this study, we analyzed the methylproteomes of both 5-fluorouracil (5-Fu) resistant Bel/5-Fu cell line and its parental Bel cell line by employing SPE-SCX based label-free quantitative proteomics. We identified 313 methylation forms on 294 sites in Bel cells and 294 methylation forms on 260 sites in Bel/5-Fu cells with high localization confidence. In addition, we quantified 251 methylation forms and found that 77 methylation forms significantly changed. After normalizing with the protein abundance, the 89 methylation forms were determined with the significant changes in site stoichiometry. The sequence characteristics of these significantly changed methylation sites are different. Gene ontology analysis showed that these significantly changed methylated proteins mainly involved in the biological processes of translation and transcription. Together, our findings indicated that protein methylation occurring in hepatocellular carcinoma might play a critical role in requiring drug resistance.Significance: The drug resistance acquired in cancer cells has been considered as a major challenge for the cancer treatment. Due to complexity, the molecular mechanisms are still largely unknown. Identifying the key markers will improve our understanding of the mechanisms and is crucial for the development of new therapeutic strategies to overcome resistance. To date, increasing number of proteomics and phosphoproteomics studies were reported to investigate the mechanisms of drug resistance. However, the methylproteomics studies related to drug resistance were not reported yet. Here, we performed the SPE-SCX based label-free quantitative proteomics to analyze the methylproteomes of both resistant cell line Bel/5-Fu and sensitive cell line Bel. Through the qualitative and quantitative analysis, we found that the sequence characteristics of methylation sites were evidently different between these two cell lines. The results suggested that some methyltransferases might play a crucial role in the regulation of drug resistance. We also performed the analysis of methyl-site stoichiometry by normalizing the protein abundances. It was found that 89 methylation forms were determined with the significant changes in site stoichiometry, which may contribute to the development of the Bel cells into resistant cells. Our methylproteomes dataset would be useful to reveal novel molecular mechanisms of drug resistance acquired in hepatocellular carcinoma.