Kawasaki H, Moriguchi T, Matsuda S, Li H Z, Nakamura S, Shimohama S, Kimura J, Gotoh Y, Nishida E
Department of Genetics and Molecular Biology, Kyoto University, Japan.
Eur J Biochem. 1996 Oct 15;241(2):315-21. doi: 10.1111/j.1432-1033.1996.00315.x.
We have previously shown that osmotic stress activates both the mitogen-activated protein kinase (MAPK) cascade and the stress-activated protein kinase (SAPK, also known as JNK) cascade in rat fibroblastic 3Y1 cells and rat PC12 cells. Here, we show that treatment of these cells with sodium arsenite, a chemical compound that mimics the effects of heat shock, or anisomycin, a protein synthesis inhibitor, induces activation of SAPKs potently. These chemical compounds also stimulated the activity of SEK1/MKK4/JNKK, SAPK activator, and the activity of MEKK, SEK1 activator. Expression of a dominant negative mutant of Ras blocked the anisomycin-induced activation of SAPK and SEK1, but did not affect markedly the arsenite-induced or heat shock-induced activation in PC12 cells. The osmotic-stress-induced activation of SAPK was insensitive to the expression of a dominant negative Ras, but was partly sensitive to down-regulation of protein kinase C. These results suggest the existence of Ras-dependent and Ras-independent activation pathways for the SAPK cascade triggered by environmental stresses including chemical stress in PC12 cells. Cell staining with a specific anti-SAPK serum showed that SAPKs were present in both the cytoplasm and the nucleus under normal conditions, and became located mainly in the nucleus after osmotic stress or ultraviolet treatment, suggesting the nuclear translocation of SAPKs.
我们之前已经表明，渗透应激可激活大鼠成纤维细胞3Y1和大鼠PC12细胞中的丝裂原活化蛋白激酶（MAPK）级联反应以及应激激活蛋白激酶（SAPK，也称为JNK）级联反应。在此，我们表明，用亚砷酸钠（一种模拟热休克效应的化合物）或茴香霉素（一种蛋白质合成抑制剂）处理这些细胞，可有效诱导SAPKs的激活。这些化合物还刺激了SAPK激活剂SEK1/MKK4/JNKK的活性以及SEK1激活剂MEKK的活性。Ras显性负性突变体的表达阻断了茴香霉素诱导的SAPK和SEK1的激活，但对PC12细胞中亚砷酸钠诱导的或热休克诱导的激活没有明显影响。渗透应激诱导的SAPK激活对显性负性Ras的表达不敏感，但对蛋白激酶C的下调部分敏感。这些结果表明，在PC12细胞中，由包括化学应激在内的环境应激触发的SAPK级联反应存在Ras依赖性和Ras非依赖性激活途径。用特异性抗SAPK血清进行细胞染色显示，在正常条件下，SAPKs存在于细胞质和细胞核中，在渗透应激或紫外线处理后主要定位于细胞核，提示SAPKs发生了核转位。
