Department of Molecular Genetics & Cellular Microbiology, Institute of Molecular Biosciences, Goethe University, Max-von-Laue Str. 9, 60438 Frankfurt/M, Germany.
Nucleic Acids Res. 2013 Oct;41(19):9062-76. doi: 10.1093/nar/gkt679. Epub 2013 Aug 2.
Yeast 25S rRNA was reported to contain a single cytosine methylation (m(5)C). In the present study using a combination of RP-HPLC, mung bean nuclease assay and rRNA mutagenesis, we discovered that instead of one, yeast contains two m(5)C residues at position 2278 and 2870. Furthermore, we identified and characterized two putative methyltransferases, Rcm1 and Nop2 to be responsible for these two cytosine methylations, respectively. Both proteins are highly conserved, which correlates with the presence of two m(5)C residues at identical positions in higher eukaryotes, including humans. The human homolog of yeast Nop2, p120 has been discovered to be upregulated in various cancer tissues, whereas the human homolog of Rcm1, NSUN5 is completely deleted in the William's-Beuren Syndrome. The substrates and function of both human homologs remained unknown. In the present study, we also provide insights into the significance of these two m(5)C residues. The loss of m(5)C2278 results in anisomycin hypersensitivity, whereas the loss of m(5)C2870 affects ribosome synthesis and processing. Establishing the locations and enzymes in yeast will not only help identifying the function of their homologs in higher organisms, but will also enable understanding the role of these modifications in ribosome function and architecture.
酵母 25S rRNA 被报道含有一个胞嘧啶甲基化(m(5)C)。在本研究中，我们使用反相高效液相色谱法、绿豆核酸酶测定法和 rRNA 诱变，发现酵母中含有两个 m(5)C 残基，分别位于位置 2278 和 2870。此外，我们鉴定并表征了两个假定的甲基转移酶 Rcm1 和 Nop2，它们分别负责这两个胞嘧啶甲基化。这两种蛋白质高度保守，这与包括人类在内的高等真核生物中在相同位置存在两个 m(5)C 残基有关。酵母 Nop2 的人类同源物 p120 已被发现在上皮细胞癌等多种癌症组织中上调，而 Rcm1 的人类同源物 NSUN5 在威廉姆斯-比伦综合征中完全缺失。这两种人类同源物的底物和功能仍然未知。在本研究中，我们还深入探讨了这两个 m(5)C 残基的意义。m(5)C2278 的缺失导致anisomycin 敏感性增加，而 m(5)C2870 的缺失则影响核糖体的合成和加工。确定酵母中的位置和酶不仅有助于识别其在高等生物中的同源物的功能，还能理解这些修饰在核糖体功能和结构中的作用。
