Kunnath L, Locker J
Department of Pathology, The University of Chicago, IL 60637, USA.
EMBO J. 1983;2(3):317-24. doi: 10.1002/j.1460-2075.1983.tb01425.x.
We have analyzed methylation of the rat albumin and alpha-fetoprotein (AFP) genes by hydridizing labeled cDNA clones to HpaII and MspI digests of DNA from different stages of development. These CCGG-cutting enzymes distinguish 5-methylcystosine in mCCGG (sensitive to HpaII) and CmCGG (sensitive to MspI). In the liver, the albumin gene is heavily methylated at 18 days gestation and uniformly demethylated in the adult. The AFP gene is also heavily methylated at 18 days gestation, and develops demethylated regions at the 3' half of the gene in the adult. These methylation changes are not observed in other embryonic or adult tissues. We also evaluated expression of these genes by measuring their corresponding mRNAs. The albumin gene is actively transcribed in 18-day fetal liver, when it is heavily methylated, as well as in adult liver, when it is unmethylated. In contrast, the AFP gene is transcribed only in fetal liver, even though it is less methylated in adult liver. These findings suggest that specific methylation changes are associated with changes in gene expression, but that this association is not adequately described by the simple hypothesis that methylation turns genes off.
我们通过将标记的cDNA克隆与来自不同发育阶段的DNA经HpaII和MspI酶切后的片段杂交，分析了大鼠白蛋白和甲胎蛋白（AFP）基因的甲基化情况。这些能切割CCGG的酶可区分mCCGG中的5-甲基胞嘧啶（对HpaII敏感）和CmCGG中的5-甲基胞嘧啶（对MspI敏感）。在肝脏中，白蛋白基因在妊娠18天时高度甲基化，在成年时则完全去甲基化。AFP基因在妊娠18天时也高度甲基化，在成年时基因的3'端一半区域出现去甲基化。在其他胚胎或成年组织中未观察到这些甲基化变化。我们还通过测量相应的mRNA评估了这些基因的表达情况。白蛋白基因在妊娠18天的胎儿肝脏中高度甲基化时仍能活跃转录，在成年肝脏中去甲基化时也能活跃转录。相反，AFP基因仅在胎儿肝脏中转录，即便它在成年肝脏中的甲基化程度较低。这些发现表明特定的甲基化变化与基因表达的变化相关，但甲基化使基因失活这一简单假说并不能充分解释这种关联。
