Fernandez-Peralta A M, Navarro P, Tagarro I, Gonzalez-Aguilera J J
Departamento de Biologia, Facultad de Ciencias, Universidad Autónoma de Madrid, Spain.
Genome. 1994 Oct;37(5):770-4. doi: 10.1139/g94-110.
The isoschizomers MspI and HpaII are four base cutter (C decrease CGG) restriction endonucleases, HpaII being sensitive to methylation of the internal cytosine. Human chromosomes treated with MspI have produced inconsistent results between laboratories, while HpaII has always been described as a nonbanding enzyme when used on human chromosomes. These results have been explained on the basis of both rarity of the CpG doublet in vertebrate genomes and high rate of CpG methylation (5mCpG). We demonstrated consistent banding patterns subsequent to digestions with MspI and HpaII. On euchromatin, MspI (but not HpaII) digests the DNA of R regions and thus R-bands apparently contain many more CCGG sites (mostly methylated) than G-bands. In heterochromatin, extensive digestion of the 9q12 region not only by MspI but also by HpaII reveals a heterochromatic domain with a high frequency of unmethylated CCGG sites, most probably within the satellite 3 DNA fraction. In addition, enzymatic digestions of the Yq12 heterochromatin, when this region is undercondensed by 5-azacytidine, contribute to deepen the insight into the mechanism of action of this cytidine analog and at the same time reinforce the idea of the heterogeneity of this chromosome region where domains with unmethylated CCGG sites may also exist.
同裂酶MspI和HpaII是四碱基切割酶（C↓CGG）限制性内切核酸酶，HpaII对内部胞嘧啶的甲基化敏感。用MspI处理人类染色体，各实验室得到的结果并不一致，而HpaII用于人类染色体时一直被描述为无带型酶。基于脊椎动物基因组中CpG二联体的稀少性和CpG甲基化（5mCpG）的高发生率，这些结果得到了解释。我们证明了用MspI和HpaII消化后有一致的带型模式。在常染色质上，MspI（而非HpaII）消化R区域的DNA，因此R带显然比G带含有更多的CCGG位点（大多为甲基化的）。在异染色质中，9q12区域不仅被MspI广泛消化，也被HpaII广泛消化，揭示出一个具有高频未甲基化CCGG位点的异染色质结构域，很可能在卫星3 DNA部分内。此外，当Yq12异染色质被5-氮杂胞苷处理而凝缩不足时，对其进行酶切消化有助于深入了解这种胞苷类似物的作用机制，同时强化了这一染色体区域存在未甲基化CCGG位点结构域的异质性观点。
