An Exploration of RNA Modification, Structure and Functionality of tRNA-Derived Small RNAs

tRNA-derived small RNAs (tsRNAs) are a type of widely existed small non-coding RNA with their mechanisms of action little explored. tsRNAs are sophisticatedly modified, creating a challenge to study their biological function because synthetic tsRNAs may not mimic their structure and interacting potential. Here, we show examples how endogenous tsRNAs with extensive RNA modifications can have cellular functions different from synthetic ones that without modifications. By adding site-specific RNA modifications to engineer synthetic tsRNAs, we mimic the endogenous tsRNAs structure conformation and recapitulate the functional effects of endogenous tsRNAs. In addition, we applied a LC-MS based de novo RNA sequencing method (MLC-seq) to profile RNA modification of endogenous tRNAs or tsRNAs. Our efforts may guide future understanding and designing of modified tsRNAs for precise gene regulation with a comprehensive tsRNA modification profile identified