Bellas R E, Bendori R, Farmer S R
Department of Biochemistry, Boston University School of Medicine, Massachusetts.
J Biol Chem. 1991 Jun 25;266(18):12008-14.
Growth activation of quiescent Swiss 3T3 fibroblasts leads to a rapid induction of vinculin and beta 1-integrin gene expression. Addition of serum, epidermal growth factor (EGF), or platelet-derived growth factor to serum-starved, density-arrested cells resulted in a rapid increase in vinculin and beta 1-integrin mRNA levels and a corresponding increase in vinculin synthesis. The increase in vinculin and beta 1-integrin mRNA expression by serum or EGF was not blocked by the inhibition of protein synthesis by cycloheximide. The kinetics of induction of vinculin and beta 1-integrin mRNAs by EGF are different: vinculin mRNA levels reached a peak of expression 4-5-fold greater than that measured in quiescent cells by 2 h after addition of growth factor, whereas beta 1-integrin mRNA levels increased more slowly and to a lesser extent, reaching peaks of 2-3-fold induction at 5 h poststimulation. Down-regulation of protein kinase C by prolonged pretreatment of cells with phorbol 1,2-myristate 1,3-acetate had no effect on the ability of EGF or platelet-derived growth factor to activate vinculin or beta 1-integrin mRNA expression. Furthermore, direct activation of protein kinase C with 1,2-myristate 1,3-acetate did not induce the expression of vinculin or beta 1-integrin mRNA, but did activate c-fos expression. In vitro nuclear "run-on" transcription assays demonstrate a greater than 7-fold increase in vinculin and beta 1-integrin transcription at 40-60 min after addition of EGF when compared with levels in quiescent cells. This activation was rapid and transient, but appeared to occur later than the increase in c-fos and actin transcription. These results demonstrate that vinculin and beta 1-integrin, important components of the cell adhesion apparatus, are members of a group of immediate early growth-responsive genes, along with c-fos, c-myc, actin, and fibronectin. In addition, regulation of these cell adhesion genes occurs exclusively through a protein kinase C-independent pathway in serum-deprived, density-arrested Swiss 3T3 cells.
静止的瑞士3T3成纤维细胞的生长激活导致纽蛋白和β1整合素基因表达的快速诱导。向血清饥饿、密度停滞的细胞中添加血清、表皮生长因子（EGF）或血小板衍生生长因子，会导致纽蛋白和β1整合素mRNA水平迅速增加，同时纽蛋白合成相应增加。血清或EGF诱导的纽蛋白和β1整合素mRNA表达增加不受放线菌酮抑制蛋白质合成的影响。EGF诱导纽蛋白和β1整合素mRNA的动力学不同：添加生长因子后2小时，纽蛋白mRNA水平达到的表达峰值比静止细胞中测量值高4 - 5倍，而β1整合素mRNA水平增加较慢且幅度较小，在刺激后5小时达到2 - 3倍诱导的峰值。用佛波醇1,2 - 十四烷酸酯1,3 - 乙酸酯对细胞进行长时间预处理来下调蛋白激酶C，对EGF或血小板衍生生长因子激活纽蛋白或β1整合素mRNA表达的能力没有影响。此外，用1,2 - 十四烷酸酯1,3 - 乙酸酯直接激活蛋白激酶C不会诱导纽蛋白或β1整合素mRNA的表达，但会激活c - fos表达。体外细胞核“连续”转录分析表明，与静止细胞中的水平相比，添加EGF后40 - 60分钟，纽蛋白和β1整合素转录增加超过7倍。这种激活迅速且短暂，但似乎比c - fos和肌动蛋白转录的增加发生得晚。这些结果表明，纽蛋白和β1整合素作为细胞粘附装置的重要组成部分，与c - fos、c - myc、肌动蛋白和纤连蛋白一样，是一组立即早期生长反应基因的成员。此外，在血清剥夺、密度停滞的瑞士3T3细胞中，这些细胞粘附基因的调控完全通过一条不依赖蛋白激酶C的途径进行。
