Differential regulation of HSC70, HSP70, HSP90 alpha, and HSP90 beta mRNA expression by mitogen activation and heat shock in human lymphocytes

A subset of heat shock proteins, HSP90 alpha, HSP90 beta, and a member of the HSP70 family, HSC70, shows enhanced synthesis following mitogenic activation as well as heat shock in human peripheral blood mononuclear cells. In this study, we have examined expression of mRNA for these proteins, including the major 70-kDa heat shock protein, HSP70, in mononuclear cells following either heat shock or mitogenic activation with phytohemagglutinin (PHA), ionomycin, and the phorbol ester, tetradecanoyl phorbol acetate. The results demonstrate that the kinetics of mRNA expression of these four genes generally parallel the kinetics of enhanced protein synthesis seen following either heat shock or mitogen activation and provide clear evidence that mitogen-induced synthesis of HSC70 and HSP90 is due to increased mRNA levels and not simply to enhanced translation of preexisting mRNA. Although most previous studies have focused on cell cycle regulation of HSP70 mRNA, we found that HSP70 mRNA was only slightly and transiently induced by PHA activation, while HSC70 is the predominant 70-kDa heat shock protein homologue induced by mitogens. Similarly, HSP90 alpha appears more inducible by heat shock than mitogens while the opposite is true for HSP90 beta. These results suggest that, although HSP70 and HSC70 have been shown to contain similar promoter regions, additional regulatory mechanisms which result in differential expression to a given stimulus must exist. They clearly demonstrate that human lymphocytes are an important model system for determining mechanisms for regulation of heat shock protein synthesis in unstressed cells. Finally, based on kinetics of mRNA expression, the results are consistent with the hypothesis that HSC70 and HSP90 gene expression are driven by an IL-2/IL-2 receptor-dependent pathway in human T cells.     

Modulation of the level of expression of cellular genes in adenovirus 12-infected and transformed human cells.

The level of expression of thymidine kinase (TK), heat shock protein 70 (HSP70), beta-tubulin and p53 was assessed in human embryo kidney cells (HEKs) infected with adenovirus type 12 (Ad 12) and Ad 12 early region 1 (E1) mutants. HSP70, beta-tubulin and p53 levels were unchanged but TK activity was dramatically increased following wild-type infection. The initial activation of TK required the expression of the product of the E1A 13S mRNA but sustained expression only occurred with those viruses expressing the E1B proteins as well. A number of human cell lines transformed with either Ad 12 or Ad 5 E1 DNA were also assessed for the level of expression of HSP70, beta-tubulin and p53. Both HSP70 and beta-tubulin levels were greatly increased compared with primary human cells although there was considerable variation between lines. p53 was only expressed at high levels in Ad 12-transformed lines expressing E1A and E1B proteins.     

腾冲嗜热厌氧菌热休克蛋白60及其基因表达的热激动态变化分析

热休克蛋白60(HSP60)是细菌体内一种非常重要的分子伴侣,其可以协助蛋白质或肽链的正确折叠和构型,防止变性和降解。基于本实验室的早期观察,腾冲嗜热厌氧菌的HSP60是一个典型的温度相关蛋白质,在80℃的表达水平最高。为了进一步了解嗜热菌应急的分子机制,继续进行了在热激后HSP60基因表达的动态研究。将最适温度(75℃)下培养的腾冲嗜热厌氧菌迅速地转移至80℃继续培养,然后在不同的时间点上分别取样,并通过双向电泳、Western blot和Real_time PCR等方法,分析了HSP60在mRNA和蛋白质水平上的表达量的改变。试验结果表明,在80℃热处理4h内的短期应急过程中,HSP60蛋白水平一直呈上升趋势,而它的mRNA水平则表现为先升高后下降的一个非对称性的峰形变化。HSP60的mRNA和蛋白质的对温度的应答快慢程度是不同的。HSP60的mRNA水平的显著变化在1h内便可观察到,而蛋白质水平的显著改变要延迟3h左右。此外,HSP60的mRNA和蛋白质对温度的应答量变大小也是不同的。