论文部分内容阅读
目的 研究丝裂原活化蛋白激酶 - (MAPK)信号传导途径对 Jurkat细胞中热休克蛋白 90基因(hsp90 )表达的影响。方法 用 Western免疫印迹 -增强型化学发光系统 (ECL)检测热休克前后 Jurkat细胞胞外信号调节激酶 (ERK)、p38激酶及 c- Jun N-末端蛋白激酶 (JNK)的底物 c- Jun的磷酸化程度 ;分别用 JNK1的显性负突变质粒 DN- JNK1、p38c DNA反义表达质粒 anti- p38及 ERK激酶的特异性抑制剂 PD980 5 9瞬时转染或处理Jurkat细胞 ,再行 RT- PCR检测 hsp90 α和 hsp90 β基因的表达水平。分别以转染空载体 (p CDNA3)及未经任何处理的 Jurkat细胞作为相应对照组。结果 45℃热休克 15 min后 ,Jurkat细胞中 JNK1和 p38激酶的磷酸化程度与热休克前相比明显增强 ,ERK的磷酸化较热休克前有所增强。转染 DN- JNK1的细胞中 hsp90 α和 hsp90 β基因的组成性和热诱导表达均较转染空载体的对照组降低 ,尤其对 hsp90 α基因的热诱导表达的影响更为明显 (仅为对照组的6 8% ) ;转染 anti- p38后 ,hsp90 α基因的组成性表达和 hsp90 β基因热诱导表达分别比转染空载体的对照组增加2 6 %和 2 2 % ;经 PD980 5 9处理的 Jurkat细胞中 ,hsp90 α基因的组成性表达比未处理的细胞增加 46 % ,但 PD980 5 9处理对 hsp90 α和 hsp90
Objective To investigate the effect of mitogen-activated protein kinase (MAPK) signaling pathway on the expression of heat shock protein 90 (hsp90) gene in Jurkat cells. Methods Western blotting-enhanced chemiluminescence (ECL) was used to detect the expression of extracellular signal-regulated kinase (ERK), p38 kinase and c-Jun of c-Jun N-terminal protein kinase (JNK) in Jurkat cells before and after heat shock Phosphorylation of Jurkat cells were transiently transfected or treated with JNK1 dominant negative mutant plasmid DN-JNK1, p38c DNA antisense plasmid anti-p38 and ERK kinase-specific inhibitor PD98059 respectively, and then RT-PCR The expression levels of hsp90α and hsp90βgene were detected. Respectively transfected with empty vector (pCDNA3) and untreated Jurkat cells as the corresponding control group. Results After 45 ℃ heat shock for 15 min, the phosphorylation of JNK1 and p38 kinase in Jurkat cells was significantly increased compared with that before heat shock. The phosphorylation of ERK was enhanced before heat shock. The constitutive and heat-induced expression of hsp90α and hsp90β genes in DN-JNK1 transfected cells were lower than those transfected with empty vector, especially the heat-induced expression of hsp90α gene (only control Group). After transfection with anti-p38, the constitutive expression of hsp90 α gene and the heat-induced expression of hsp90 β gene were increased by 26% and 22% respectively compared with the control group transfected with empty vector. After PD9805 9 In Jurkat cells treated, the constitutive expression of the hsp90α gene increased by 46% compared to untreated cells, but treatment with PD98059 on hsp90α and hsp90