Drug tolerance requires switching from low to high OXPHOS in AML cells. (A-B) Flow cytometry (A) and quantification (B) of OP-puro incorporation into nascent polypeptides in mitochondria of TRMT5-depleted (sg2, sg4) and control (sgCtr) MOLM13 cells. Puromycin-treated cells served as a control. (C-D) OCR in OCI-AML2 (C) and MOLM13 (D) TRMT5 KO (sg2, sg4) and sgCtr cells. (E-F) OCR in drug-tolerant OCI-AML2, OCI-AML3 (F) and drug-sensitive MOLM13 and Kasumi-1 (F) cells exposed to cytarabine (AraC) (orange) or untreated (control) (blue). (G-J) OCR in rescued TRMT5-KO cells (sg2) using the WT, enzymatic dead MUT, mitochondria-deficient (ΔMTS) constructs of TRMT5, or the EV as control in the presence (+AraC) or absence of cytarabine. (K) Cum FRQ of log₂ FC of nascent protein abundance of mitochondrial proteins (MitoCarta3.0; n = 182) in AraC-treated control (scr) OCI-AML2 cells (yellow), AraC-treated TRMT5-KO cells (red), or AraC TRMT5-KO cells vs control (scr) cells (blue). Scr: Infection of a scramble sgRNA. (L-N) Correlation of log₂ FC of nascent translation in TRMT5 OCI-AML2 (OCI2) and MOLM13-KO cells in the absence (L) or presence (M) of AraC and corresponding gene enrichment analyses (N) (GOrilla). Mean ± standard deviation (panels B-J). Dunnett multiple comparison test (panel B). Wilcoxon matched-pair signed rank test (panel K). ATP, adenosine triphosphate; ADP, adenosine 5′-diphosphate; Cum FRQ, cumulative frequency; FCCP, carbonylcyanide-p-trifluoromethoxyphenylhydrazone; FDR, false discovery rate; GO, Gene Ontology; OP-puro, O-propargyl-puromycin; Rot/AA, rotenone and antimycin A.