ABSTRACT

Aminoacyl-tRNA synthetases (aaRSs) are a conserved family of enzymes with an essential role in protein synthesis: ligating amino acids to cognate tRNA molecules for translation. In addition to their role in tRNA charging, aaRSs have acquired non-canonical functions, including post-transcriptional regulation of mRNA expression. Yet, the extent and mechanisms of these post-transcriptional functions are largely unknown. Herein, we performed a comprehensive transcriptome analysis to define the mRNAs that are associated with almost all aaRSs present in S. cerevisiae cytosol. Nineteen (out of twenty) isogenic strains of GFP-tagged cytosolic aaRSs were subjected to immunoprecipitation with anti-GFP beads along with an untagged control. mRNAs associated with each aaRS were then identified by RNA-seq. The extent of mRNA association varied significantly between aaRSs, from MetRS in which none appeared to be statistically significant, to PheRS that binds hundreds of different mRNAs. Interestingly, many target mRNAs are bound by multiple aaRSs, suggesting co-regulation by this family of enzymes. Gene Ontology analyses for aaRSs with a considerable number of target mRNAs discovered an enrichment for pathways of amino acid metabolism and of ribosome biosynthesis. Furthermore, sequence and structure motif analysis revealed for some aaRSs an enrichment for motifs that resemble the anticodon stem loop of cognate tRNAs. These data suggest that aaRSs coordinate mRNA expression in response to amino acid availability and may utilize RNA elements that mimic their canonical tRNA binding partners.

 抽象的

氨酰基-tRNA 合成酶 (aaRS) 是一个保守的酶家族，在蛋白质合成中发挥重要作用：将氨基酸连接到同源 tRNA 分子以进行翻译。除了在 tRNA 充电中的作用外，aaRS 还获得了非规范功能，包括 mRNA 表达的转录后调节。然而，这些转录后功能的程度和机制在很大程度上尚不清楚。在此，我们进行了全面的转录组分析，以确定与酿酒酵母胞质中存在的几乎所有 aaRS 相关的 mRNA。使用抗 GFP 珠子和未标记的对照对 19 个（共 20 个）等基因菌株的 GFP 标记的胞质 aaRS 进行免疫沉淀。然后通过 RNA-seq 鉴定与每个 aaRS 相关的 mRNA。 aaRS 之间的 mRNA 关联程度差异显着，从 MetRS（其中没有一个似乎具有统计学意义）到 PheRS（结合数百种不同的 mRNA）。有趣的是，许多靶标 mRNA 与多个 aaRS 结合，表明该酶家族的共同调节。对具有大量目标 mRNA 的 aaRS 进行基因本体分析，发现氨基酸代谢和核糖体生物合成途径富集。此外，序列和结构基序分析揭示了一些 aaRS 的基序富集，类似于同源 tRNA 的反密码子茎环。这些数据表明，aaRS 响应氨基酸可用性来协调 mRNA 表达，并可能利用模仿其典型 tRNA 结合伴侣的 RNA 元件。