Department of Biochemistry and Translational Research Center for Protein Function Control, Yonsei University, Seoul 120-749, Korea.
J Biol Chem. 2010 Dec 31;285(53):41280-9. doi: 10.1074/jbc.M110.182188. Epub 2010 Nov 2.
Osmotic stress causes profound perturbations of cell functions. Although the adaptive responses required for cell survival upon osmotic stress are being unraveled, little is known about the effects of osmotic stress on ubiquitin-dependent proteolysis. We now report that hyperosmotic stress inhibits proteasome activity by activating p38 MAPK. Osmotic stress increased the level of polyubiquitinated proteins in the cell. The selective p38 inhibitor SB202190 decreased osmotic stress-associated accumulation of polyubiquitinated proteins, indicating that p38 MAPK plays an inhibitory role in the ubiquitin proteasome system. Activated p38 MAPK stabilized various substrates of the proteasome and increased polyubiquitinated proteins. Proteasome preparations purified from cells expressing activated p38 MAPK had substantially lower peptidase activities than control proteasome samples. Proteasome phosphorylation sites dependent on p38 were identified by measuring changes in the extent of proteasome phosphorylation in response to p38 MAPK activation. The residue Thr-273 of Rpn2 is the major phosphorylation site affected by p38 MAPK. The mutation T273A in Rpn2 blocked the proteasome inhibition that is mediated by p38 MAPK. These results suggest that p38 MAPK negatively regulates the proteasome activity by phosphorylating Thr-273 of Rpn2.
渗透胁迫会对细胞功能造成严重干扰。尽管人们正在揭示细胞在渗透胁迫下生存所需的适应性反应，但对于渗透胁迫对泛素依赖性蛋白水解的影响知之甚少。我们现在报告，高渗胁迫通过激活 p38 MAPK 来抑制蛋白酶体活性。渗透胁迫会增加细胞内多泛素化蛋白的水平。选择性 p38 抑制剂 SB202190 减少了与渗透胁迫相关的多泛素化蛋白的积累，表明 p38 MAPK 在泛素蛋白酶体系统中起抑制作用。激活的 p38 MAPK 稳定了蛋白酶体的各种底物，并增加了多泛素化蛋白的含量。与对照蛋白酶体样品相比，从表达激活的 p38 MAPK 的细胞中纯化的蛋白酶体制剂的肽酶活性明显降低。通过测量蛋白酶体磷酸化程度对 p38 MAPK 激活的反应变化，确定了依赖于 p38 的蛋白酶体磷酸化位点。Rpn2 的残基 Thr-273 是受 p38 MAPK 影响的主要磷酸化位点。Rpn2 中的 T273A 突变阻断了由 p38 MAPK 介导的蛋白酶体抑制。这些结果表明，p38 MAPK 通过磷酸化 Rpn2 的 Thr-273 来负调控蛋白酶体活性。
