López-Boado Y S, Tolivia J, López-Otín C
Departamento de Biología Funcional, Universidad de Oviedo, Spain.
J Biol Chem. 1994 Oct 28;269(43):26871-8.
We have examined the regulation by retinoic acid of the gene encoding apolipoprotein D (apoD), a human plasma protein belonging to the superfamily of the lipocalins that is produced by a specific subtype of highly differentiated breast carcinomas. Northern blot analysis revealed that all-trans-retinoic acid (RA) strongly induced the accumulation of apoD mRNA in T-47D and ZR-75-1 estrogen receptor-positive human breast cancer cells in a time- and dose-dependent manner, while no inductive effect was observed in estrogen receptor-negative cell lines, including MDA-MB-231 and MDA-MB-435. The effect of RA on apoD expression by T-47D cells was at least 12-fold more potent than the effect of the steroids dihydrotestosterone and dexamethasone, which had been previously described as hormonal up-regulators of apoD expression in these cells. A time course study demonstrated that the induction of apoD mRNA reached a level of 15-fold over the untreated control cells after 48 h of incubation in the presence of a 10(-7) M concentration of RA. A dose-response analysis showed that as little as 10(-13) M RA produced an accumulation of 5-fold over the control, while incubation of the cells in the presence of 10(-5) M RA induced a maximal accumulation of 24-fold over the control untreated cells. The induction of apoD mRNA was independent of the synthesis of proteins de novo, as demonstrated by the fact that the induction was also detected in the presence of cycloheximide. The incubation of the cells in the presence of RA did not affect significantly the stability of apoD mRNA. By contrast, treatment of the T-47D cells with RA produced an increase of approximately 8-fold in the rate of transcription of the apoD gene. Furthermore, treatment of the T-47D cells with RA induced the synthesis and secretion to the culture medium of apoD. This increased expression of apoD was accompanied by an inhibition of cell proliferation and a progression through a more differentiated phenotype, suggesting that the mechanisms controlling RA-induced growth arrest, cell differentiation, and apoD synthesis may be directly coordinated in human breast cancer cells.
我们研究了视黄酸对载脂蛋白D（apoD）编码基因的调控作用。apoD是一种人类血浆蛋白，属于脂质运载蛋白超家族，由高分化乳腺癌的一种特定亚型产生。Northern印迹分析显示，全反式视黄酸（RA）以时间和剂量依赖性方式强烈诱导T-47D和ZR-75-1雌激素受体阳性人乳腺癌细胞中apoD mRNA的积累，而在包括MDA-MB-231和MDA-MB-435在内的雌激素受体阴性细胞系中未观察到诱导作用。RA对T-47D细胞中apoD表达的作用比类固醇二氢睾酮和地塞米松的作用至少强12倍，而类固醇二氢睾酮和地塞米松此前被描述为这些细胞中apoD表达的激素上调因子。一项时间进程研究表明，在存在10^(-7) M浓度RA的情况下孵育48小时后，apoD mRNA的诱导水平比未处理的对照细胞高出15倍。剂量反应分析表明，低至10^(-13) M的RA产生的积累比对照高出5倍，而在存在10^(-5) M RA的情况下孵育细胞诱导的积累比未处理的对照细胞最大高出24倍。apoD mRNA的诱导与从头合成蛋白质无关，这一事实表明在存在放线菌酮的情况下也能检测到诱导作用。在存在RA的情况下孵育细胞对apoD mRNA的稳定性没有显著影响。相比之下，用RA处理T-47D细胞使apoD基因的转录速率增加了约8倍。此外，用RA处理T-47D细胞诱导了apoD的合成并分泌到培养基中。apoD表达的这种增加伴随着细胞增殖的抑制和向更分化表型的进展，这表明在人乳腺癌细胞中控制RA诱导的生长停滞、细胞分化和apoD合成的机制可能直接相互协调。
