TRMT6/61A-dependent base methylation of tRNA-derived fragments regulates gene-silencing activity and the unfolded protein response in bladder cancer

RNA modifications are important regulatory elements of RNA functions. However, most genome-wide mapping of RNA modifications has focused on messenger RNAs and transfer RNAs, but such datasets have been lacking for small RNAs. Here we mapped N(1)-methyladenosine (m(1)A) in the cellular small RNA space. Benchmarked with synthetic m(1)A RNAs, our workflow identified specific groups of m(1)A-containing small RNAs, which are otherwise disproportionally under-represented. In particular, 22-nucleotides long 3′ tRNA-fragments are highly enriched for TRMT6/61A-dependent m(1)A located within the seed region. TRMT6/61A-dependent m(1)A negatively affects gene silencing by tRF-3s. In urothelial carcinoma of the bladder, where TRMT6/61A is over-expressed, higher m(1)A modification on tRFs is detected, correlated with a dysregulation of tRF targetome. Lastly, TRMT6/61A regulates tRF-3 targets involved in unfolded protein response. Together, our results reveal a mechanism of regulating gene expression via base modification of small RNA.