Wan Y J, Pan T, Wang L, Locker J, Wu T C
Department of Pathology, Harbor-UCLA Medical Center, Torrance 90509, USA.
J Mol Endocrinol. 1995 Feb;14(1):101-8. doi: 10.1677/jme.0.0140101.
In McA-RH 8994 rat hepatoma cells, all-trans-retinoic acid (t-RA) induces expression of the alpha-fetoprotein (AFP) and albumin genes and results in a phenotype similar to differentiated fetal hepatocytes. The present study elucidated the mechanism involved in AFP gene regulation mediated by retinoic acid. Northern blot analyses demonstrated that 9-cis-retinoic acid (c-RA), a ligand for retinoid x receptors (RXRs), also induced expression of the AFP gene in McA-RH 8994 cells. The induction was time- and dose-dependent. Northern blots and transfection assays using the 7.3 kb full-length regulatory region of the AFP gene demonstrated that c-RA was more effective than t-RA in regulating expression of the AFP gene. At 10(-7) M, c-RA increased AFP mRNA 5-fold and chloramphenicol acetyltransferase (CAT) activity 2.5-fold. In contrast, t-RA at a concentration of 10(-7) M exerted no significant effect; 10(-6) to 10(-5) M t-RA was needed to affect AFP gene expression. These data suggested that activation of RXRs is essential for the regulation of the AFP gene. Co-transfection experiments revealed that over-expression of RXR alpha in McA-RH 8994 cells further enhanced the CAT activity induced by c-RA. In addition, c-RA did not alter the half-life of AFP mRNA. Thus, RXR alpha may play a crucial role in transcriptional regulation of the AFP gene and in controlling hepatocyte phenotype.
在McA-RH 8994大鼠肝癌细胞中，全反式维甲酸（t-RA）可诱导甲胎蛋白（AFP）和白蛋白基因的表达，并产生一种类似于分化胎儿肝细胞的表型。本研究阐明了维甲酸介导的AFP基因调控机制。Northern印迹分析表明，9-顺式维甲酸（c-RA）作为维甲酸X受体（RXR）的配体，也可在McA-RH 8994细胞中诱导AFP基因的表达。这种诱导具有时间和剂量依赖性。使用AFP基因7.3 kb全长调控区进行的Northern印迹和转染分析表明，c-RA在调控AFP基因表达方面比t-RA更有效。在10^(-7) M浓度下，c-RA可使AFP mRNA增加5倍，氯霉素乙酰转移酶（CAT）活性增加2.5倍。相比之下，10^(-7) M浓度的t-RA没有显著作用；需要10^(-6)至10^(-5) M的t-RA才能影响AFP基因表达。这些数据表明，RXR的激活对于AFP基因的调控至关重要。共转染实验表明，在McA-RH 8994细胞中过表达RXRα可进一步增强c-RA诱导的CAT活性。此外，c-RA并未改变AFP mRNA的半衰期。因此，RXRα可能在AFP基因的转录调控以及控制肝细胞表型方面发挥关键作用。
