Nakhasi H L, Lynch K R, Dolan K P, Unterman R, Antakly T, Feigelson P
J Biol Chem. 1982 Mar 25;257(6):2726-9.
alpha 2u globulin synthesis ceases when a liver cell becomes malignant. We have compared the structure and transcription of the alpha 2u globulin genes in Morris hepatomas 5123D and 7793 with that of normal hepatic genes using alpha 2u globulin cDNA as a hybridization probe. No alpha 2u globulin mRNA was detected in hepatoma 7793 by cell-free translation, Northern blot, or R0t analysis. In hepatoma 5123D, however, a small number of alpha 2u globulin RNA sequences were detected by Northern blot and R0t analysis, but this RNA was not detectably translationally active either in vivo or in an in vitro cell-free translational system. No structural differences between normal liver and the hepatoma alpha 2u globulin genes were observed by Southern blot analysis of genomic DNA digested with restriction endonucleases Pst I, Bam HI, or Ava I. However, a general demethylation of cytosine residues in the alpha 2u globulin genes of hepatomas has been found using the restriction enzyme Hha I and the isoschizomeric pair of restriction enzymes Msp I and Hpa II. Therefore, loss of the alpha 2u globulin phenotype in these hepatomas is accompanied by extensive demethylation of DNA sequences within and/or immediately flanking the alpha 2u globulin genes.
当肝细胞发生癌变时，α2u球蛋白的合成会停止。我们使用α2u球蛋白cDNA作为杂交探针，比较了莫里斯肝癌5123D和7793中α2u球蛋白基因的结构和转录情况与正常肝脏基因的差异。通过无细胞翻译、Northern印迹或R0t分析，在肝癌7793中未检测到α2u球蛋白mRNA。然而，在肝癌5123D中，通过Northern印迹和R0t分析检测到了少量的α2u球蛋白RNA序列，但这种RNA在体内或体外无细胞翻译系统中均未检测到明显的翻译活性。用限制性内切酶Pst I、Bam HI或Ava I消化基因组DNA后进行Southern印迹分析，未观察到正常肝脏和肝癌α2u球蛋白基因之间的结构差异。然而，使用限制性内切酶Hha I以及同裂酶对Msp I和Hpa II，发现肝癌α2u球蛋白基因中的胞嘧啶残基普遍发生了去甲基化。因此，这些肝癌中α2u球蛋白表型的丧失伴随着α2u球蛋白基因内部和/或紧邻其侧翼的DNA序列的广泛去甲基化。
