A CRISPR dropout screen for tRNA regulators identified YRDC as the top essential gene in glioblastoma stem cells. Threonine acts as a substrate of YRDC to facilitate the N6-threonylcarbamoyladenosine (t6A) tRNA modification and shift translation toward mitosis-related genes with a codon bias. Our findings support targeting glioblastoma growth by a well-tolerated dietary therapy.