Messeguer R, Ganal M W, Steffens J C, Tanksley S D
Department of Plant Breeding and Biometry, Cornell University, Ithaca, NY 14853.
Plant Mol Biol. 1991 May;16(5):753-70. doi: 10.1007/BF00015069.
The tomato nuclear genome was determined to have a G + C content of 37% which is among the lowest reported for any plant species. Non-coding regions have a G + C content even lower (32% average) whereas coding regions are considerably richer in G + C (46%). 5-methyl cytosine was the only modified base detected and on average 23% of the cytosine residues are methylated. Immature tissues and protoplasts have significantly lower levels of cytosine methylation (average 20%) than mature tissues (average 25%). Mature pollen has an intermediate level of methylation (22%). Seeds gave the highest value (27%), suggesting de novo methylation after pollination and during seed development. Based on isoschizomer studies we estimate 55% of the CpG target sites (detected by Msp I/Hpa II) and 85% of the CpNpG target sites (detected by Bst NI/Eco RI) are methylated. Unmethylated target sites (both CpG and CpNpG) are not randomly distributed throughout the genome, but frequently occur in clusters. These clusters resemble CpG islands recently reported in maize and tobacco. The low G + C content and high levels of cytosine methylation in tomato may be due to previous transitions of 5mC----T. This is supported by the fact that G + C levels are lowest in non-coding portions of the genome in which selection is relaxed and thus transitions are more likely to be tolerated. This hypothesis is also supported by the general deficiency of methylation target sites in the tomato genome, especially in non-coding regions. Using methylation isoschizomers and RFLP analysis we have also determined that polymorphism between plants, for cytosine methylation at allelic sites, is common in tomato. Comparing DNA from two tomato species, 20% of the polymorphisms detected by Bst NI/Eco RII could be attributed to differential methylation at the CpNpG target sites. With Msp I/Hpa II, 50% of the polymorphisms were attributable to methylation (CpG and CpNpG sites). Moreover, these polymorphisms were demonstrated to be inherited in a mendelian fashion and to co-segregate with the methylation target site and thus do not represent variation for transacting factors that might be involved in methylation of DNA. The potential role of heritable methylation polymorphism in evolution of gene regulation and in RFLP studies is discussed.
番茄核基因组的G + C含量被测定为37%，这在已报道的所有植物物种中是最低的之一。非编码区的G + C含量更低（平均32%），而编码区的G + C含量则高得多（46%）。5-甲基胞嘧啶是唯一检测到的修饰碱基，平均23%的胞嘧啶残基被甲基化。未成熟组织和原生质体的胞嘧啶甲基化水平（平均20%）显著低于成熟组织（平均25%）。成熟花粉的甲基化水平处于中间值（22%）。种子的甲基化水平最高（27%），这表明授粉后及种子发育过程中发生了从头甲基化。基于同裂酶研究，我们估计55%的CpG靶位点（由Msp I/Hpa II检测）和85%的CpNpG靶位点（由Bst NI/Eco RI检测）被甲基化。未甲基化的靶位点（包括CpG和CpNpG）并非随机分布于整个基因组，而是经常成簇出现。这些簇类似于最近在玉米和烟草中报道的CpG岛。番茄中低G + C含量和高水平的胞嘧啶甲基化可能是由于先前5mC向T的转变。基因组非编码部分的G + C水平最低，在这些区域选择压力较小，因此这种转变更可能被容忍，这一事实支持了这一观点。番茄基因组中甲基化靶位点普遍缺乏，尤其是在非编码区，这也支持了这一假说。利用甲基化同裂酶和RFLP分析，我们还确定了番茄中植物间等位基因位点胞嘧啶甲基化的多态性很常见。比较两个番茄物种的DNA，Bst NI/Eco RII检测到的20%的多态性可归因于CpNpG靶位点的差异甲基化。对于Msp I/Hpa II，50%的多态性可归因于甲基化（CpG和CpNpG位点）。此外，这些多态性被证明以孟德尔方式遗传，并与甲基化靶位点共分离，因此不代表可能参与DNA甲基化的反式作用因子的变异。本文讨论了可遗传的甲基化多态性在基因调控进化和RFLP研究中的潜在作用。
