Viard I, Wehrli P, Jornot L, Bullani R, Vechietti J L, Schifferli J A, Tschopp J, French L E
Department of Dermatology, Geneva University Medical School, Switzerland.
J Invest Dermatol. 1999 Mar;112(3):290-6. doi: 10.1046/j.1523-1747.1999.00531.x.
Clusterin is a widely expressed, well conserved, secreted glycoprotein, which is highly induced in tissues regressing as a consequence of apoptotic cell death in vivo. It has recently been shown that clusterin expression is only confined to surviving cells following the induction of apoptosis in vitro, suggesting that it is involved in cell survival rather than death. In the hypothesis that clusterin may be implicated in cellular responses to stress, clusterin gene expression was analyzed in the A431 human epidermoid cancer cell line following heat shock and oxidative stress. Our results show that both a transient heat shock (20 min at 42 degrees C) and various oxidative stresses, including hydrogen peroxide, superoxide anion, hyperoxia and UVA exposure, induce a strong increase in clusterin mRNA levels as assessed by northern blot. Nuclear run-on analysis suggests that transcriptional activation is involved in inducing clusterin mRNA in response to heat shock. Using pulse-chase analysis of control and heat shocked cells, it is shown that clusterin mRNA is translated and secreted, thus resulting in increased extracellular levels of the protein following heat shock. To investigate the function of clusterin in response to these stresses, clusterin anti-sense transfectants that stably express virtually no clusterin at the mRNA and protein level were generated in A431 cells. These anti-sense transfectants are shown to be highly sensitive to apoptotic cell death induced by heat shock or oxidative stress compared with wild-type A431 cells or control transfectants. Taken together, our results show that clusterin gene expression is induced in response to heat shock and oxidative stress in human A431 cells, and confers cellular protection against heat shock and oxidative stress.
聚集素是一种广泛表达、高度保守的分泌型糖蛋白，在体内因凋亡性细胞死亡而发生退化的组织中高度诱导表达。最近研究表明，在体外诱导细胞凋亡后，聚集素的表达仅局限于存活细胞，这表明它参与细胞存活而非死亡。基于聚集素可能参与细胞应激反应的假说，我们分析了热休克和氧化应激后A431人表皮样癌细胞系中聚集素基因的表达。我们的结果显示，短暂热休克（42℃，20分钟）以及包括过氧化氢、超氧阴离子、高氧和紫外线A照射在内的各种氧化应激，通过Northern印迹法评估，均可诱导聚集素mRNA水平显著升高。核转录分析表明，转录激活参与热休克诱导聚集素mRNA的过程。通过对对照细胞和热休克细胞进行脉冲追踪分析，发现聚集素mRNA被翻译并分泌，因此热休克后细胞外该蛋白水平升高。为了研究聚集素在应对这些应激时的功能，我们在A431细胞中构建了稳定表达几乎没有mRNA和蛋白质水平聚集素的反义转染细胞系。与野生型A431细胞或对照转染细胞相比，这些反义转染细胞系对热休克或氧化应激诱导的凋亡性细胞死亡高度敏感。综上所述，我们的结果表明，在人A431细胞中，热休克和氧化应激可诱导聚集素基因表达，并赋予细胞对热休克和氧化应激的保护作用。
