Protein C-terminal enzymatic labeling identifies novelcaspase cleavages during the apoptosis of multiple myeloma cells induced bykinase inhibition
Protein C-terminal enzymatic labeling identifies novelcaspase cleavages during the apoptosis of multiple myeloma cells induced bykinase inhibition
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蛋白 C 末端酶标记可识别激酶抑制诱导的多发性骨髓瘤细胞凋亡过程中的新型半胱天冬酶裂解
DOI：
10.1002/pmic.201500356
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发表时间：
2016
期刊：
影响因子：
3.4
通讯作者：
Guoqiang Xu
中科院分区：
文献类型：
--
作者：
Wenwen Duan;Suping Chen;Yang Zhang;Dan Li;Rong Wang;Shi Chen;Junbei Li;Xiaoyan Qiu;Guoqiang Xu
Caspase的激活和蛋白水解裂解是细胞凋亡早期的主要事件。通过对半胱天冬酶切割的蛋白底物的鉴定，将揭示细胞凋亡过程中早期事件的发生，并可能为癌症治疗提供潜在的药物靶点。虽然已经开发了几种基于N末端MS的蛋白质组学方法来鉴定蛋白水解裂解，但这些方法具有其固有的缺点。在这里，我们应用先前开发的蛋白质组学方法，蛋白质C末端酶标记（ProC-TEL），以确定在激酶抑制诱导的骨髓瘤细胞系凋亡的早期阶段发生的caspase裂解事件。检测到先前鉴定的和新的半胱天冬酶切割位点，并在激酶抑制后通过生物化学证实了几种蛋白质表达水平的降低，尽管目前的ProC-TEL程序尚未完全优化，无法提供与N-末端标记方法相当的肽鉴定。所鉴定的裂解蛋白形成复杂的相互作用网络，其中中心枢纽决定细胞凋亡期间的形态学变化。序列分析表明，当使用传统的N末端组学方法时，一些ProC-TEL鉴定的半胱天冬酶切割事件是无法鉴定的。这项工作表明，ProC‐TEL是N‐末端组学的一种补充方法，用于鉴定蛋白水解裂解事件，如信号通路中的半胱天冬酶裂解。
Caspase activation and proteolytic cleavages are the major events in the early stage of apoptosis. Identification of protein substrates cleaved by caspases will reveal the occurrence of the early events in the apoptotic process and may provide potential drug targets for cancer therapy. Although several N‐terminal MS‐based proteomic approaches have been developed to identify proteolytic cleavages, these methods have their inherent drawbacks. Here we apply a previously developed proteomic approach, protein C‐terminal enzymatic labeling (ProC‐TEL), to identify caspase cleavage events occurring in the early stage of the apoptosis of a myeloma cell line induced by kinase inhibition. Both previously identified and novel caspase cleavage sites are detected and the reduction of the expression level of several proteins is confirmed biochemically upon kinase inhibition although the current ProC‐TEL procedure is not fully optimized to provide peptide identifications comparable to N‐terminal labeling approaches. The identified cleaved proteins form a complex interaction network with central hubs determining morphological changes during the apoptosis. Sequence analyses show that some ProC‐TEL identified caspase cleavage events are unidentifiable when traditional N‐terminomic approaches are utilized. This work demonstrates that ProC‐TEL is a complementary approach to the N‐terminomics for the identification of proteolytic cleavage events such as caspase cleavages in signaling pathways.