Faculty of Biology, Technion-Israel Institute of Technology, Haifa 3200003, Israel.
Department of Biological Chemistry, Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190401, Israel.
Int J Mol Sci. 2023 Aug 4;24(15):12442. doi: 10.3390/ijms241512442.
The p38 members of the mitogen-activated protein kinases (MAPKs) family mediate various cellular responses to stress conditions, inflammatory signals, and differentiation factors. They are constitutively active in chronic inflammatory diseases and some cancers. The differences between their transient effects in response to signals and the chronic effect in diseases are not known. The family is composed of four isoforms, of which p38α seems to be abnormally activated in diseases. p38α and p38β are almost identical in sequence, structure, and biochemical and pharmacological properties, and the specific unique effects of each of them, if any, have not yet been revealed. This study aimed to reveal the specific effects induced by p38α and p38β, both when transiently activated in response to stress and when chronically active. This was achieved via large-scale proteomics and phosphoproteomics analyses using stable isotope labeling of two experimental systems: one, mouse embryonic fibroblasts (MEFs) deficient in each of these p38 kinases and harboring either an empty vector or vectors expressing p38α, p38β, or intrinsically active variants of these MAPKs; second, induction of transient stress by exposure of MEFs, p38α, and p38β MEFs to anisomycin. Significant differences in the repertoire of the proteome and phosphoproteome between cells expressing active p38α and p38β suggest distinct roles for each kinase. Interestingly, in both cases, the constitutive activation induced adaptations of the cells to the chronic activity so that known substrates of p38 were downregulated. Within the dramatic effect of p38s on the proteome and phosphoproteome, some interesting affected phosphorylation sites were those found in cancer-associated p53 and Hspb1 (HSP27) proteins and in cytoskeleton-associated proteins. Among these, was the stronger direct phosphorylation by p38α of p53-Ser309, which was validated on the Ser315 in human p53. In summary, this study sheds new light on the differences between chronic and transient p38α and p38β signaling and on the specific targets of these two kinases.
丝裂原活化蛋白激酶（MAPK）家族的 p38 成员介导各种细胞对应激条件、炎症信号和分化因子的反应。它们在慢性炎症性疾病和一些癌症中持续活跃。其对信号的短暂作用与疾病中的慢性作用之间的差异尚不清楚。该家族由四个同工型组成，其中 p38α 在疾病中似乎异常激活。p38α 和 p38β 在序列、结构、生化和药理学特性上几乎相同，它们各自的特定独特作用，如果有的话，尚未揭示。本研究旨在揭示 p38α 和 p38β 在应激时短暂激活和慢性激活时诱导的特定效应。这是通过两种实验系统的大规模蛋白质组学和磷酸蛋白质组学分析来实现的：一种是缺乏这些 p38 激酶之一的小鼠胚胎成纤维细胞（MEF），并且携带空载体或表达 p38α、p38β 或这些 MAPKs 的固有活性变体的载体；其次，通过使 MEF、p38α 和 p38β MEF 暴露于anisomycin 来诱导瞬时应激。表达活性 p38α 和 p38β 的细胞中蛋白质组和磷酸蛋白质组的明显差异表明每种激酶的作用不同。有趣的是，在这两种情况下，慢性活性诱导的细胞适应性导致 p38 的已知底物下调。在 p38 对蛋白质组和磷酸蛋白质组的强烈影响中，一些有趣的受影响磷酸化位点是在癌症相关的 p53 和 Hspb1（HSP27）蛋白和细胞骨架相关蛋白中发现的。其中包括 p38α 对 p53-Ser309 的更强直接磷酸化，这在人类 p53 中的 Ser315 上得到了验证。总之，本研究为慢性和短暂的 p38α 和 p38β 信号之间的差异以及这两种激酶的特定靶点提供了新的认识。
