Department of Cell Biology, Harvard Medical School, Boston, MA, USA.
Nat Struct Mol Biol. 2019 May;26(5):343-349. doi: 10.1038/s41594-019-0211-4. Epub 2019 Apr 22.
Aberrantly stalled ribosomes initiate the ribosome-associated quality control (RQC) and mRNA surveillance pathways for the degradation of potentially toxic peptides and faulty mRNAs. During RQC, ANKZF1 (yeast Vms1p) releases ubiquitinated nascent proteins from 60S ribosomal subunits for proteasomal degradation. Here, we use a cell-free system to show that ANKZF1 and Vms1p sever polypeptidyl-tRNAs on RQC complexes by precisely cleaving off the terminal 3'CCA nucleotides universal to all tRNAs. This produces a tRNA fragment that cannot be aminoacylated until its 3'CCA end is restored. The recycling of ANKZF1-cleaved tRNAs is intact in the mammalian cytosol via a two-step process that requires the removal of a 2',3'-cyclic phosphate and TRNT1, the sole CCA-adding enzyme that mediates tRNA biogenesis in eukaryotes. TRNT1 also discriminates between properly folded tRNA substrates and aberrant tRNA substrates, selectively tagging the latter for degradation. Thus, ANKZF1 liberates peptidyl-tRNAs from stalled ribosomes such that the tRNA is checked in an obligate way for integrity before reentry into the translation cycle.
异常停滞的核糖体启动核糖体相关的质量控制（RQC）和 mRNA 监测途径，以降解潜在毒性肽和有缺陷的 mRNAs。在 RQC 过程中，ANKZF1（酵母 Vms1p）将泛素化的新生蛋白从 60S 核糖体亚基上释放出来，进行蛋白酶体降解。在这里，我们使用无细胞系统表明，ANKZF1 和 Vms1p 通过精确地切断所有 tRNA 通用的末端 3'CCA 核苷酸，在 RQC 复合物上切断多肽-tRNAs。这产生了一个 tRNA 片段，直到其 3'CCA 末端恢复之前，不能被氨酰化。ANKZF1 切割的 tRNA 通过两步过程在哺乳动物细胞质中完整循环，这需要去除 2'，3'-环磷酸和 TRNT1，后者是唯一的 CCA-添加酶，介导真核生物的 tRNA 生物发生。TRNT1 还区分正常折叠的 tRNA 底物和异常的 tRNA 底物，选择性地标记后者进行降解。因此，ANKZF1 将肽-tRNAs 从停滞的核糖体中释放出来，使得 tRNA 在重新进入翻译循环之前以强制性的方式检查其完整性。
