Graduate School of Life Science, University of Hyogo, Ako-gun 678-1297, Japan.
Graduate School of Materials Science, Nagoya University, Nagoya 464-8602, Japan.
Nucleic Acids Res. 2019 Jun 20;47(11):5936-5949. doi: 10.1093/nar/gkz270.
In eukaryotes and archaea, tRNA genes frequently contain introns, which are removed during maturation. However, biological roles of tRNA introns remain elusive. Here, we constructed a complete set of Saccharomyces cerevisiae strains in which the introns were removed from all the synonymous genes encoding 10 different tRNA species. All the intronless strains were viable, but the tRNAPheGAA and tRNATyrGUA intronless strains displayed slow growth, cold sensitivity and defective growth under respiratory conditions, indicating physiological importance of certain tRNA introns. Northern analyses revealed that removal of the introns from genes encoding three tRNAs reduced the amounts of the corresponding mature tRNAs, while it did not affect aminoacylation. Unexpectedly, the tRNALeuCAA intronless strain showed reduced 5.8S rRNA levels and abnormal nucleolar morphology. Because pseudouridine (Ψ) occurs at position 34 of the tRNAIleUAU anticodon in an intron-dependent manner, tRNAIleUAU in the intronless strain lost Ψ34. However, in a portion of tRNAIleUAU population, position 34 was converted into 5-carbamoylmethyluridine (ncm5U), which could reduce decoding fidelity. In summary, our results demonstrate that, while introns are dispensable for cell viability, some introns have diverse roles, such as ensuring proper growth under various conditions and controlling the appropriate anticodon modifications for accurate pairing with the codon.
在真核生物和古菌中，tRNA 基因经常包含内含子，这些内含子在成熟过程中被切除。然而，tRNA 内含子的生物学功能仍然难以捉摸。在这里，我们构建了一套完整的酿酒酵母菌株，其中所有编码 10 种不同 tRNA 物种的同义基因中的内含子都被切除。所有无内含子的菌株都是可行的，但 tRNAPheGAA 和 tRNATyrGUA 无内含子的菌株表现出生长缓慢、对冷敏感以及在呼吸条件下生长缺陷，表明某些 tRNA 内含子具有生理重要性。Northern 分析显示，从编码三种 tRNA 的基因中去除内含子会降低相应成熟 tRNA 的数量，而不会影响氨酰化。出乎意料的是，tRNALeuCAA 无内含子的菌株表现出 5.8S rRNA 水平降低和核仁形态异常。因为假尿嘧啶 (Ψ) 在 tRNAIleUAU 反密码子的第 34 位以依赖内含子的方式存在，所以在无内含子的菌株中，tRNAIleUAU 失去了 Ψ34。然而，在 tRNAIleUAU 群体的一部分中，第 34 位被转化为 5-羧甲基尿嘧啶 (ncm5U)，这可能会降低解码保真度。总之，我们的结果表明，虽然内含子对于细胞活力不是必需的，但有些内含子具有多种功能，例如确保在各种条件下的适当生长，以及控制适当的反密码子修饰以与密码子进行准确配对。
