Yeast Nfs1p is involved in thio-modification of both mitochondrial and cytoplasmic tRNAs
Yeast Nfs1p is involved in thio-modification of both mitochondrial and cytoplasmic tRNAs
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DOI：
10.1074/jbc.m312448200
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发表时间：
2004-03-26
影响因子：
4.8
通讯作者：
Kagamiyama, H
中科院分区：
文献类型：
--
作者：
Nakai, Y;Umeda, N;Kagamiyama, H
IscS蛋白是一种含磷酸吡哆醛的半胱氨酸脱硫酶，参与铁硫簇生物合成。在原核生物中，IscS还参与各种代谢功能，包括tRNA的硫代修饰。相比之下，IscS（Nfs 1）的真核直系同源物迄今已被证明仅在线粒体铁硫簇生物发生中起作用。我们在这里证明，酵母Nfs 1 p也需要在体内的线粒体（mt）和细胞质（cy）tRNA的转录后硫代修饰。Nfs 1 p的缺失导致mt-tRNA（UUU）（Lys）和mt-tRNA（UUG）（Gln）摆动位置上5-羧甲基氨基甲基-2-硫代尿苷的2-硫代修饰立即受损。此外，我们观察到cy-tRNA（UUU）（Lys 2）和cy-tRNA（UUC）（Glu 3）的5-甲氧羰基甲基-2-硫代尿苷（mcm（5）s（2）U）的2-硫代修饰严重减少，尽管与mt-tRNA相比，这种效应有些延迟。质谱分析显示，从Nfs 1 p缺失的细胞中制备的cy-tRNA中，5-甲氧羰基甲基尿苷增加，同时mcm（5）s（2）U减少。这些结果表明，Nfs 1 p参与了mt-tRNA中5-羧甲基氨基甲基-2-硫代尿苷和cy-tRNA中mcm（5）s（2）U的2-硫代修饰。
The IscS protein is a pyridoxal phosphate-containing cysteine desulfurase involved in iron-sulfur cluster biogenesis. In prokaryotes, IscS is also involved in various metabolic functions, including thio-modification of tRNA. By contrast, the eukaryotic ortholog of IscS (Nfs1) has thus far been shown to be functional only in mitochondrial iron-sulfur cluster biogenesis. We demonstrate here that yeast Nfs1p is also required for the post-transcriptional thio-modification of both mitochondrial (mt) and cytoplasmic (cy) tRNAs in vivo. Depletion of Nfs1p resulted in an immediate impairment of the 2-thio-modification of 5-carboxymethylaminomethyl-2-thiouridine at the wobble positions of mt-tRNA(UUU)(Lys) and mt-tRNA(UUG)(Gln). In addition, we observed a severe reduction in the 2-thiomodification of 5-methoxycarbonylmethyl-2-thiouridine (mcm(5)s(2)U) of cy-tRNA(UUU)(Lys2) and cy-tRNA(UUC)(Glu3), although the effect was somewhat delayed compared with that seen in mt-tRNAs. Mass spectrometry analysis revealed an increase in 5-methoxycarbonylmethyluridine concomitant with a decrease in mcm(5)s(2)U in cy-tRNAs that were prepared from Nfs1p-depleted cells. These results suggest that Nfs1p is involved in the 2-thio-modification of both 5-carboxymethylaminomethyl-2-thiouridine in mt-tRNAs and mcm(5)s(2)U in cy-tRNAs.