Lin K H, Brennan M D, Lindahl R
Department of Biology, University of Alabama, Tuscaloosa 35487.
Cancer Res. 1988 Dec 15;48(24 Pt 1):7009-12.
Significant changes in aldehyde dehydrogenase (ALDH) activity occur during rat hepatocarcinogenesis in vivo. To compare the structure and expression of the tumor aldehyde dehydrogenase gene in rat hepatoma cell lines and normal rat liver, several rat hepatoma cell lines, including HTC, H4-II-EC3, JM2, McA-RH7777, and four lines established in this laboratory have been examined for T-ALDH gene expression using a tumor ALDH complementary DNA. Northern blot analysis of polyadenylate-containing RNA from log-phase cells and normal rat liver with T-ALDH complementary DNA indicates production of a single major 1.7-kilobase transcript in the high activity lines HTC, JM2, RLT-2M, RLT-3C, RLT-9F, and intermediate activity line RLT-5G. There is a direct correlation between expression of T-ALDH enzyme activity and the amount of 1.7-kilobase transcript. S1 nuclease protection experiments confirm that there is only one major T-ALDH transcript in the high activity lines. Thus, cell line differences in T-ALDH activity are reflected in the level of a single T-ALDH transcript. Southern analysis was used to identify the T-ALDH gene in genomic DNA. The results indicate that no significant amplification or rearrangement of the T-ALDH gene has occurred in these hepatoma cells. DNA methylation has been proposed to play an important role in gene expression. Genomic DNA from HTC, JM2, McA-RH7777, H4-II-EC3, RLT-2M, RLT-9F, RLT-3C, RLT-5G, rat embryo and normal rat liver were digested with MspI and HpaII to examine methylation patterns. A digestion pattern consistent with hypomethylation was detected only in DNA from the high T-ALDH activity cell lines HTC, JM2, RLT-2M, and RLT-9F. This suggests that constitutive expression of T-ALDH in the hepatoma cells is related to changes in DNA methylation patterns.
在大鼠体内肝癌发生过程中，醛脱氢酶（ALDH）活性会发生显著变化。为比较大鼠肝癌细胞系和正常大鼠肝脏中肿瘤醛脱氢酶基因的结构与表达，我们使用肿瘤ALDH互补DNA，对几种大鼠肝癌细胞系进行了检测，其中包括HTC、H4-II-EC3、JM2、McA-RH7777，以及本实验室建立的四个细胞系。用T-ALDH互补DNA对对数期细胞和正常大鼠肝脏中含多聚腺苷酸的RNA进行Northern印迹分析，结果表明，在高活性细胞系HTC、JM2、RLT-2M、RLT-3C、RLT-9F和中等活性细胞系RLT-5G中，产生了一条主要的1.7千碱基转录本。T-ALDH酶活性的表达与1.7千碱基转录本的量之间存在直接相关性。S1核酸酶保护实验证实，高活性细胞系中只有一条主要的T-ALDH转录本。因此，T-ALDH活性的细胞系差异反映在单一T-ALDH转录本的水平上。Southern分析用于鉴定基因组DNA中的T-ALDH基因。结果表明，这些肝癌细胞中未发生T-ALDH基因的显著扩增或重排。有人提出DNA甲基化在基因表达中起重要作用。用MspI和HpaII消化HTC、JM2、McA-RH7777、H4-II-EC3、RLT-2M、RLT-9F、RLT-3C、RLT-5G、大鼠胚胎和正常大鼠肝脏的基因组DNA，以检测甲基化模式。仅在高T-ALDH活性细胞系HTC、JM2、RLT-2M和RLT-9F的DNA中检测到与低甲基化一致的消化模式。这表明肝癌细胞中T-ALDH的组成型表达与DNA甲基化模式的变化有关。
