BOT-02 2-METHYLTHIO MODIFICATION OF N6-ISOPENTENYLADENOSINE IN MITOCHONDRIAL TRNAS BY CDK5RAP1 PROMOTES THE MAINTENANCE OF GLIOMA-INITIATING CELLS

2-Methylthio-N(6)-isopentenyl modification of adenosine (ms(2)i(6)A) is an evolutionally conserved modification that is found in mitochondrial (mt)-tRNAs. Cdk5 regulatory subunit-associated protein 1 (CDK5RAP1) specifically converts N6-isopentenyladenosine (i(6)A) to ms(2)i(6)A at position A37 of four mt-DNA-encoded tRNAs, and the modification regulates efficient mitochondrial translation and energy metabolism in mammals. Here, we report that the ms(2) conversion mediated by CDK5RAP1 in mt-tRNAs is required to sustain glioma-initiating cell (GIC)-related traits. CDK5RAP1 maintained the self-renewal capacity, undifferentiated state, and tumorigenic potential of GICs. This regulation was not related to the translational control of mt-proteins. CDK5RAP1 abrogated the antitumor effect of i(6)A by converting i(6)A to ms(2)i(6)A and protected GICs from excessive autophagy triggered by i(6)A. The elevated activity of CDK5RAP1 contributed to the amelioration of the cytotoxic effect of i(6)A and promoted GIC maintenance. The hypoxic microenvironment in the tumor core activated CDK5RAP1, whose activity was inversely correlated with the oxygen concentration because of two [4Fe-4S] clusters in the enzyme. This work demonstrates that CDK5RAP1 is crucial for the detoxification of endogenous i(6)A and that GICs readily utilize this mechanism for survival.