Department of Epigenetics and Molecular Carcinogenesis, University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA.
Department of Medical Microbiology and Immunology, and Program in Bioinformatics, The University of Toledo College of Medicine and Life Sciences, Toledo, OH 43614, USA.
Nucleic Acids Res. 2020 Oct 9;48(18):10329-10341. doi: 10.1093/nar/gkaa604.
The recently characterized mammalian writer (methyltransferase) and eraser (demethylase) of the DNA N6-methyladenine (N6mA) methyl mark act on single-stranded (ss) and transiently-unpaired DNA. As YTH domain-containing proteins bind N6mA-containing RNA in mammalian cells, we investigated whether mammalian YTH domains are also methyl mark readers of N6mA DNA. Here, we show that the YTH domain of YTHDC1 (known to localize in the nucleus) binds ssDNA containing N6mA, with a 10 nM dissociation constant. This binding is stronger by a factor of 5 than in an RNA context, tested under the same conditions. However, the YTH domains of YTHDF2 and YTHDF1 (predominantly cytoplasmic) exhibited the opposite effect with ∼1.5-2× stronger binding to ssRNA containing N6mA than to the corresponding DNA. We determined two structures of the YTH domain of YTHDC1 in complex with N6mA-containing ssDNA, which illustrated that YTHDC1 binds the methylated adenine in a single-stranded region flanked by duplexed DNA. We discuss the hypothesis that the writer-reader-eraser of N6mA-containining ssDNA is associated with maintaining genome stability. Structural comparison of YTH and SRA domains (the latter a DNA 5-methylcytosine reader) revealed them to be diverse members of a larger family of DNA/RNA modification readers, apparently having originated from bacterial modification-dependent restriction enzymes.
最近发现的哺乳动物 DNA N6-甲基腺嘌呤（N6mA）甲基标记的书写者（甲基转移酶）和橡皮擦（去甲基酶）作用于单链（ss）和瞬时未配对的 DNA。由于 YTH 结构域蛋白结合含有 N6mA 的 RNA 存在于哺乳动物细胞中，我们研究了哺乳动物 YTH 结构域是否也是 N6mA DNA 甲基标记的读取器。在这里，我们表明 YTHDC1 的 YTH 结构域（已知定位于细胞核中）与含有 N6mA 的 ssDNA 结合，解离常数为 10 nM。在相同条件下进行测试时，这种结合比在 RNA 环境中要强 5 倍。然而，YTHDF2 和 YTHDF1 的 YTH 结构域（主要存在于细胞质中）表现出相反的效果，与相应的 DNA 相比，对含有 N6mA 的 ssRNA 的结合要强约 1.5-2 倍。我们确定了 YTHDC1 的 YTH 结构域与含有 N6mA 的 ssDNA 复合物的两个结构，这说明了 YTHDC1 以单链区域结合被修饰的腺嘌呤，该单链区域由双链 DNA 侧翼。我们讨论了这样一个假设，即含有 N6mA 的 ssDNA 的书写者-读取器-橡皮擦与维持基因组稳定性有关。YTH 和 SRA 结构域（后者是 DNA 5-甲基胞嘧啶的读取器）的结构比较表明，它们是一个更大的 DNA/RNA 修饰读取器家族的不同成员，显然源自细菌依赖修饰的限制酶。
