Non-lethal stress-induced discrete proteolysis is caspase-3/7 dependent

Cells are in constant adaptation to environmental changes to insure their proper functioning. When exposed to stresses, cells activate specific pathways to elicit adaptive modifications. Those changes can be mediated by selective modulation of gene and protein expression as well as by post-translational modifications, such as phosphorylation and proteolytic processing. Protein cleavage, as a controlled and limited post-translational modification, is involved in diverse physiological processes such as the maintenance of protein homeostasis, activation of repair pathways, apoptosis and the regulation of proliferation. Here we assessed by quantitative proteomics the proteolytic landscape in two cell lines subjected to low cisplatin concentrations used as a mild non-lethal stress paradigm. This landscape was compared to the one obtained in the same cells stimulated with cisplatin concentrations inducing apoptosis. These analyses were performed in wild-type cells and in cells lacking the two main executioner caspases: caspase-3 and caspase-7. Ninety proteins were found to be cleaved at one or a few sites (discrete cleavage) in low stress conditions compared to four hundred and forty in apoptotic cells. Many of the cleaved proteins in stressed cells were also found to be cleaved in apoptotic conditions. As expected in apoptotic cells, ~80% of the cleavage events were dependent on caspase 3/caspase 7. Strikingly, upon exposure to non-lethal stresses, no discrete cleavage was detected in cells lacking caspase-3 and caspase-7. This indicates that the proteolytic landscape in stressed viable cells fully depends on the activity of executioner caspases. These results suggest that the so-called executioner caspases fulfill important stress adaptive responses distinct from their role in apoptosis.

细胞始终处于适应环境变化的状态，以保障其正常功能的运转。当受到应激刺激时，细胞会激活特定通路以引发适应性修饰。此类适应性改变可通过对基因与蛋白质表达的选择性调控，以及磷酸化、蛋白水解加工等翻译后修饰（post-translational modifications）介导实现。作为一种受调控且具有局限性的翻译后修饰类型，蛋白裂解（Protein cleavage）参与了诸多生理过程，包括蛋白质稳态维持、修复通路激活、细胞凋亡（apoptosis）以及细胞增殖调控。本研究通过定量蛋白质组学（quantitative proteomics），分析了两种经低浓度顺铂（cisplatin）处理的细胞系的蛋白裂解图谱，其中低浓度顺铂被用作轻度非致死应激模型。研究将该图谱与经高浓度顺铂诱导细胞凋亡的同款细胞所得的裂解图谱进行了对比。本次分析分别在野生型细胞以及缺失两种主要效应半胱天冬酶（caspase）——半胱天冬酶-3（caspase-3）与半胱天冬酶-7（caspase-7）——的细胞中开展。与凋亡细胞中检测到的440个裂解蛋白相比，轻度应激条件下共有90个蛋白在单个或少数位点发生裂解（离散型裂解，discrete cleavage）。应激细胞中诸多发生裂解的蛋白，在凋亡条件下同样可被检测到裂解现象。正如预期，凋亡细胞中约80%的裂解事件依赖于半胱天冬酶-3与半胱天冬酶-7的活性。值得注意的是，在暴露于非致死应激的细胞中，缺失半胱天冬酶-3与半胱天冬酶-7的细胞未检测到任何离散型裂解事件。这表明，存活的应激细胞内的蛋白裂解图谱完全依赖于效应半胱天冬酶的活性。本研究结果提示，所谓的效应半胱天冬酶还承担了与细胞凋亡功能截然不同的重要应激适应性应答功能。