Torres Adrian Gabriel, Piñeyro David, Rodríguez-Escribà Marta, Camacho Noelia, Reina Oscar, Saint-Léger Adélaïde, Filonava Liudmila, Batlle Eduard, Ribas de Pouplana Lluís
Institute for Research in Biomedicine (IRB Barcelona), C/Baldiri Reixac 10, Barcelona, 08028 Catalonia, Spain.
Institute for Research in Biomedicine (IRB Barcelona), C/Baldiri Reixac 10, Barcelona, 08028 Catalonia, Spain Catalan Institution for Research and Advanced Studies (ICREA), P/Lluis Companys 23, Barcelona, 08010 Catalonia, Spain.
Nucleic Acids Res. 2015 May 26;43(10):5145-57. doi: 10.1093/nar/gkv277. Epub 2015 Apr 27.
Transfer RNAs (tRNAs) are key adaptor molecules of the genetic code that are heavily modified post-transcriptionally. Inosine at the first residue of the anticodon (position 34; I34) is an essential widespread tRNA modification that has been poorly studied thus far. The modification in eukaryotes results from a deamination reaction of adenine that is catalyzed by the heterodimeric enzyme adenosine deaminase acting on tRNA (hetADAT), composed of two subunits: ADAT2 and ADAT3. Using high-throughput small RNA sequencing (RNAseq), we show that this modification is incorporated to human tRNAs at the precursor tRNA level and during maturation. We also functionally validated the human genes encoding for hetADAT and show that the subunits of this enzyme co-localize in nucleus in an ADAT2-dependent manner. Finally, by knocking down HsADAT2, we demonstrate that variations in the cellular levels of hetADAT will result in changes in the levels of I34 modification in all its potential substrates. Altogether, we present RNAseq as a powerful tool to study post-transcriptional tRNA modifications at the precursor tRNA level and give the first insights on the biology of I34 tRNA modification in metazoans.
转运RNA（tRNA）是遗传密码的关键衔接分子，在转录后会进行大量修饰。反密码子第一个残基（第34位；I34）处的肌苷是一种广泛存在的重要tRNA修饰，迄今为止对其研究较少。真核生物中的这种修饰是由异二聚体酶——作用于tRNA的腺苷脱氨酶（hetADAT）催化的腺嘌呤脱氨反应产生的，hetADAT由两个亚基组成：ADAT2和ADAT3。利用高通量小RNA测序（RNAseq），我们发现这种修饰在前体tRNA水平和成熟过程中被整合到人类tRNA中。我们还在功能上验证了编码hetADAT的人类基因，并表明该酶的亚基以ADAT2依赖的方式共定位于细胞核中。最后，通过敲低HsADAT2，我们证明hetADAT细胞水平的变化将导致其所有潜在底物中I34修饰水平的改变。总之，我们将RNAseq作为一种强大的工具，用于在前体tRNA水平研究转录后tRNA修饰，并首次深入了解后生动物中I34 tRNA修饰的生物学特性。
