Blackburn E H, Pan W C, Johnson C C
Nucleic Acids Res. 1983 Aug 11;11(15):5131-45. doi: 10.1093/nar/11.15.5131.
We have investigated the occurrence of methylated adenine residues in the macronuclear ribosomal RNA genes of Tetrahymena thermophila. It has been shown previously that macronuclear DNA, including the palindromic ribosomal RNA genes (rDNA), of Tetrahymena thermophila contains the modified base N-6-methyladenine, but no 5-methylcytosine. Purified rDNA was digested with restriction enzymes Sau 3AI, MboI and DpnI to map the positions and levels of N-6-methyladenine in the sequence 5' GATC 3'. A specific pattern of doubly methylated GATC sequences was found; hemimethylated sites were not detected. The patterns and levels of methylation of these sites did not change significantly in different physiological states. A molecular form of the rDNA found in the newly developing macronucleus and for several generations following the sexual process, conjugation, contained no detectably methylated GATC sites. However, both the bulk macronuclear DNA and palindromic rDNA from the same macronuclei were methylated. Possible roles for N-6-methyladenine in macronuclear DNA are discussed in light of these findings.
我们研究了嗜热四膜虫大核核糖体RNA基因中甲基化腺嘌呤残基的存在情况。先前已经表明，嗜热四膜虫的大核DNA，包括回文核糖体RNA基因（rDNA），含有修饰碱基N-6-甲基腺嘌呤，但不含5-甲基胞嘧啶。用限制性内切酶Sau 3AI、MboI和DpnI消化纯化的rDNA，以确定5' GATC 3'序列中N-6-甲基腺嘌呤的位置和水平。发现了一种特定的双甲基化GATC序列模式；未检测到半甲基化位点。这些位点的甲基化模式和水平在不同生理状态下没有显著变化。在新发育的大核以及有性过程（接合）后的几代中发现的一种rDNA分子形式，没有可检测到的甲基化GATC位点。然而，来自同一大核的大量大核DNA和回文rDNA都是甲基化的。根据这些发现讨论了N-6-甲基腺嘌呤在大核DNA中的可能作用。
