Abstract
Transfer RNAs are essential for translating genetic information into proteins. The human genome contains hundreds of predicted tRNA genes, many in multiple copies. How their expression is regulated to control tRNA repertoires is unknown. Here we combined quantitative tRNA profiling and chromatin immunoprecipitation with sequencing to measure tRNA expression following the differentiation of human induced pluripotent stem cells into neuronal and cardiac cells. We find that tRNA transcript levels vary substantially, whereas tRNA anticodon pools, which govern decoding rates, are more stable among cell types. Mechanistically, RNA polymerase III transcribes a wide range of tRNA genes in human induced pluripotent stem cells but on differentiation becomes constrained to a subset we define as housekeeping tRNAs. This shift is mediated by decreased mTORC1 signalling, which activates the RNA polymerase III repressor MAF1. Our data explain how tRNA anticodon pools are buffered to maintain decoding speed across cell types and reveal that mTORC1 drives selective tRNA expression during differentiation.