Capowski E E, Wells J M, Harrison G S, Karrer K M
Department of Biology, Brandeis University, Waltham, Massachusetts 02254.
Mol Cell Biol. 1989 Jun;9(6):2598-605. doi: 10.1128/mcb.9.6.2598-2605.1989.
We have cloned two DNA fragments containing 5'-GATC-3' sites at which the adenine is methylated in the macronucleus of the ciliate Tetrahymena thermophila. Using these cloned fragments as molecular probes, we analyzed the maintenance of methylation patterns at two partially and two uniformly methylated sites. Our results suggest that a semiconservative copying model for maintenance of methylation is not sufficient to account for the methylation patterns we found during somatic growth of Tetrahymena. Although we detected hemimethylated molecules in macronuclear DNA, they were present in both replicating and nonreplicating DNA. In addition, we observed that a complex methylation pattern including partially methylated sites was maintained during vegetative growth. This required the activity of a methylase capable of recognizing and modifying sites specified by something other than hemimethylation. We suggest that a eucaryotic maintenance methylase may be capable of discriminating between potential methylation sites to ensure the inheritance of methylation patterns.
我们克隆了两个含有5'-GATC-3'位点的DNA片段，在嗜热四膜虫的大核中腺嘌呤在这些位点发生甲基化。利用这些克隆片段作为分子探针，我们分析了两个部分甲基化位点和两个完全甲基化位点的甲基化模式的维持情况。我们的结果表明，用于维持甲基化的半保留复制模型不足以解释我们在四膜虫体细胞生长过程中发现的甲基化模式。尽管我们在大核DNA中检测到了半甲基化分子，但它们在复制和非复制DNA中均有存在。此外，我们观察到在营养生长过程中，包括部分甲基化位点在内的复杂甲基化模式得以维持。这需要一种甲基转移酶的活性，该酶能够识别并修饰由半甲基化以外的其他因素指定的位点。我们认为，一种真核生物维持甲基转移酶可能能够区分潜在的甲基化位点，以确保甲基化模式的遗传。
