Kochanek S, Toth M, Dehmel A, Renz D, Doerfler W
University for Genetics, University of Cologne, Germany.
Proc Natl Acad Sci U S A. 1990 Nov;87(22):8830-4. doi: 10.1073/pnas.87.22.8830.
The DNA in mammalian genomes is characterized by complex patterns of DNA methylation that reflect the states of all genetic activities of that genome. The modified nucleotide 5-methyldeoxycytidine (5mdC) can affect the interactions of specific proteins with DNA sequence motifs. The most extensively studied effect of sequence-specific methylations is that of the long-term silencing of eukaryotic (mammalian) promoters. We have initiated studies on the methylation status of parts of the human genome to view patterns of DNA methylation as indicators for genetic activities. In this report, analyses using both restriction enzyme--Southern blotting and the very precise genomic sequencing technique have been done. The genes for tumor necrosis factors (TNF) alpha and beta--in particular, their 5'-upstream and promoter regions--have been investigated in DNA isolated from human lymphocytes, granulocytes, and sperm. The results are characterized by a remarkable interindividual concordance of DNA methylation in specific human cell types. The patterns are identical in the DNA from one cell type for different individuals even of different genetic origins but different in the DNA from different cell types. As an example, in the DNA from human granulocytes of 15 different individuals (ages 20-48 yr, both sexes), 5mdC residues have been localized by the genomic sequencing technique in three identical sequence positions in the 5'-upstream region and in one downstream position of the gene encoding TNF-alpha. The promoter of this gene is free of 5mdC, and TNF-alpha is expressed in human granulocytes. The TNF-beta promoter is methylated in granulocytes from 9 different individuals, and TNF-beta is not expressed. In human lymphocytes, the main source of TNF-beta, the TNF-beta promoter is free of 5mdC residues. All 5'-CG-3' sites studied in the TNF-alpha and -beta genes are methylated in DNA from human sperm. In human cell lines HL-60, Jurkat, and RPMI 1788, the extent of DNA methylation in TNF-alpha and -beta genes has also been studied.
哺乳动物基因组中的DNA具有复杂的DNA甲基化模式，这些模式反映了该基因组所有遗传活动的状态。修饰的核苷酸5-甲基脱氧胞苷（5mdC）会影响特定蛋白质与DNA序列基序的相互作用。序列特异性甲基化最广泛研究的效应是真核（哺乳动物）启动子的长期沉默。我们已开始对人类基因组部分区域的甲基化状态进行研究，将DNA甲基化模式视为遗传活动的指标。在本报告中，我们使用了限制性内切酶- Southern印迹法和非常精确的基因组测序技术进行了分析。对从人类淋巴细胞、粒细胞和精子中分离出的DNA中的肿瘤坏死因子（TNF）α和β基因，特别是它们的5'上游和启动子区域进行了研究。结果的特点是特定人类细胞类型中DNA甲基化存在显著的个体间一致性。即使不同个体具有不同的遗传背景，一种细胞类型的DNA模式对于不同个体也是相同的，但不同细胞类型的DNA模式则不同。例如，在1十五个不同个体（年龄20 - 48岁，男女皆有）的人类粒细胞DNA中，通过基因组测序技术已将5mdC残基定位在编码TNF-α基因的5'上游区域的三个相同序列位置和一个下游位置。该基因的启动子不含5mdC，且TNF-α在人类粒细胞中表达。TNF-β启动子在9个不同个体的粒细胞中被甲基化，且TNF-β不表达。在人类淋巴细胞（TNF-β的主要来源）中，TNF-β启动子不含5mdC残基。在人类精子DNA中，TNF-α和-β基因中所有研究的5'-CG-3'位点均被甲基化。在人类细胞系HL-60、Jurkat和RPMI 1788中，也研究了TNF-α和-β基因中的DNA甲基化程度。
