Decades of study have revealed more than 100 ribonucleoside structures incorporated as post-transcriptional modifications mainly in
tRNA and rRNA, yet the larger functional dynamics of this conserved system are unclear. To this end, we developed a highly precise mass spectrometric method to quantify
tRNA modifications in Saccharomyces cerevisiae. Our approach revealed several novel biosynthetic pathways for
RNA modifications and led to the discovery of signature changes in the spectrum of
tRNA modifications in the damage response to mechanistically different toxicants. This is illustrated with the
RNA modifications Cm, m(5)C, and
m(2) (2)G, which increase following
hydrogen peroxide exposure but decrease or are unaffected by exposure to
methylmethane sulfonate,
arsenite, and
hypochlorite. Cytotoxic
hypersensitivity to
hydrogen peroxide is conferred by loss of
enzymes catalyzing the formation of Cm, m(5)C, and
m(2) (2)G, which demonstrates that
tRNA modifications are critical features of the cellular stress response. The results of our study support a general model of dynamic control of
tRNA modifications in cellular response pathways and add to the growing repertoire of mechanisms controlling translational responses in cells.