Ca^(2+)/CaM/CaMK调控热应激诱导小鼠胚胎成纤维细胞HSP72的表达

Ca^(2+)/CaM/CaMK regulating HSP72 expression in MEFs

细胞受应激后合成HSP72以保护细胞免受损伤,热应激时Ca^(2+)/CaM和HSP72的表达高度相关,但Ca^(2+)/CaM是否调节HSP72表达及其机制尚不清楚。本研究将从12.5日龄胚胎获得的小鼠胚胎成纤维细胞(MEFs)于39℃下进行热休克处理,并分别置于含CaCl_2、EGTA、W7和MyrAIP培养基中处理。分别采用RT-PCR和Western-Blot检测相关基因和蛋白表达。结果显示,在39℃热应激时,CaCl_2显著提高了细胞HSP72和CaM的表达,而EGTA和W7则显著降低了细胞HSP72和CaM的表达。同时,热应激上调了热激转录因子1(HSF1)、p-HSF1及HSP70蛋白的表达。然而,Myr-AIP显著抑制了细胞HSP72和p-HSF1蛋白的表达,而不影响HSF1蛋白的表达。这表明,热应激时,Ca^(2+)/CaM可通过活化CaMKⅡ诱导体外培养的MEFs细胞HSP72的表达。

Most cells, exposed to stress, would synthesize heat shock protein 72 （HSP72）, which is a molecular chaperone that protects cells from further stress damage. Many studies have shown that Ca^2＋/ CaM were highly correlated with HSP72 expressions when the cells were suffered from heat stress. How- ever, the mechanism of Ca^2＋/CAM inducing HSP72 expressions in response to heat stress was not clear. MEFs, prepared from 12.5-day-old mouse embryos, were heat-shocked at 39 %. The cells were treated with CaC12, EGTA, W7, and Myr-AIP medium, respectively. The gene and protein expressions were de- tected by real-time PCR and western-blot, respectively. The experimental results showed that CaC12 en- hanced both HSP72 and CaM mRNA expressions in MEFs whereas EGTA reduced their expressions. Treatment with W7 reduced both HSP72 and CaMKⅡ mRNA and protein expressions in the MEFs.Meanwhile, the HSF1, p-HSF1 and HSP72 protein expressions were upregulated by heat stress. Howev- er, Myr-AIP inhibited both HSP72 and p-HSF1 protein expressions, while it had no effect on HSF1 ex- pression in the heat-stressed MEFs. The above results suggested that Ca^2＋/CaM induced HSP72 expres- sion in heat-stressed MEFs by activating CaMKⅡ in vitro.