CELL AGING--GROWTH FACTOR CONTROL OF EARLY RESPONSE GENE

随着正常人类细胞的增殖寿命接近尾声 它们无法通过复制它们的基因来应对各种生长因素 DNA我们已经广泛地检测了WI-38细胞中的生长因子反应 并发现它们并不是失去必要的增长所造成的 因子受体或配体-受体相互作用中可识别的变化。 事实上，许多正常的有丝分裂原刺激细胞周期依赖的事件 在衰老的细胞中诱导活动，就像在年轻的细胞中一样 这些细胞确实通过进入S阶段来做出反应。这导致了 推测衰老细胞在G1末期附近被阻断 S阶段的开始。 最近我们和其他人(坎皮西和同事的个人交流) 已经观察到原癌基因c-fos的mrna似乎是 在有丝分裂原刺激的衰老细胞中检测不到。C-的表达 FOS是有丝分裂原刺激后非常早期的事件，达到最大值 在年轻细胞中大约0.5小时后的水平。我们随后有了 观察到c-fos实际上在衰老的WI-38细胞中表达 在血清或表皮生长因子刺激(EGF)之后，但它 只有在蛋白质合成同时被抑制的情况下才能被检测到 环己亚胺。这与不成比例的快速 这种信使核糖核酸在衰老细胞中的降解。我们打算研究 C-fos表达的调控和另外两个早期反应基因的调控， 青年和老年WI-38中c-jun与血清反应因子(SRF)的关系 细胞。我们将研究转录、蛋白质合成和 翻译后的磷酸化修饰。我们还将直接 检测c-fos能否延长c-jun细胞的增殖寿命 用含有这些基因的质粒转化年轻细胞 金属硫蛋白启动子控制下的正常基因。

As normal human cells approach the end of their proliferative lifespan they fail to respond to a variety of growth factors by replicating their DNA. We have examined growth factor responses extensively in WI-38 cells and have found that they do not result from the loss of necessary growth factor receptors or recognizable changes in ligand-receptor interactions. In fact, many of the normal mitogen stimulated cell cycle dependent events and activities are induced in senescent cells just as they are in young cells which do respond by entering the S phase. This has led to the speculation that senescent cells become blocked in late G1 near the beginning of the S phase. Recently we and others (Campisi and co-workers personal communication) have observed that the mRNA for the protooncogene c-fos appears to be undetectable in mitogen stimulated senescent cells. The expression of c- fos is a very early event following mitogen stimulation, reaching maximum levels after approximately 0.5 hr in young cells. We have subsequently observed that c-fos is actually expressed in senescent WI-38 cells following either serum or epidermal growth factor stimulation (EGF) but it is only detectable if protein synthesis is simultaneously inhibited with cycloheximide. This is consistent with the disproportionately rapid degradation of this mRNA in senescent cells. We intend to study the regulation of c-fos expression and that of two other early response genes, c-jun and the serum response factor (SRF) in young and senescent WI-38 cells. We will examine transcription, protein synthesis and posttranslational modification by phosphorylation. We will also directly test whether c-fos of c-jun can increase the proliferative life span of WI-38 cells by transforming young cells with plasmids containing these normal genes under the control of the metallothionein promotor.