The ten-eleven translocation 2 (TET2) protein, which oxidizes 5-methylcytosine in DNA, can also bind RNA; however, the targets and function of TET2–RNA interactions in vivo are not fully understood. Using stringent affinity tags introduced at the Tet2 locus, we purified and sequenced TET2-crosslinked RNAs from mouse embryonic stem cells (mESCs) and found a high enrichment for tRNAs. RNA immunoprecipitation with an antibody against 5-hydroxymethylcytosine (hm5C) recovered tRNAs that overlapped with those bound to TET2 in cells. Mass spectrometry (MS) analyses revealed that TET2 is necessary and sufficient for the deposition of the hm5C modification on tRNA. Tet2 knockout in mESCs affected the levels of several small noncoding RNAs originating from TET2-bound tRNAs that were enriched by hm5C immunoprecipitation. Thus, our results suggest a new function of TET2 in promoting the conversion of 5-methylcytosine to hm5C on tRNA and regulating the processing or stability of different classes of tRNA fragments.

10-11 易位 2 (TET2) 蛋白可氧化 DNA 中的 5-甲基胞嘧啶，也可以与 RNA 结合。然而，TET2-RNA 在体内相互作用的靶点和功能尚不完全清楚。使用在Tet2基因座处引入的严格亲和标签，我们从小鼠胚胎干细胞 (mESCs) 中纯化并测序了 TET2 交联的 RNA，并发现 tRNA 的高度富集。用针对 5-羟甲基胞嘧啶 (hm5C) 的抗体进行 RNA 免疫沉淀可回收与细胞中与 TET2 结合的 tRNA 重叠的 tRNA。质谱 (MS) 分析表明，TET2 对于 hm5C 修饰在 tRNA 上的沉积是必要且充分的。春节2mESCs 中的敲除影响了源自 TET2 结合的 tRNAs 的几个小的非编码 RNAs 的水平，这些 tRNAs 通过 hm5C 免疫沉淀富集。因此，我们的结果表明 TET2 在促进 tRNA 上 5-甲基胞嘧啶转化为 hm5C 和调节不同类别 tRNA 片段的加工或稳定性方面具有新功能。