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Y. Saito et al.
When Evi1/MF9 or WT/MF9 cells were treated with the glutaminase inhibitor BPTES, Evi1/MF9 cells (but not WT/MF9 cells) were sensitive to inhibition of OCR (Figure 4I). Collectively, these data indicate that EVI1 plays an essential role in mitochondrial energy reprogramming in MF9 AML cells by activating glutaminolysis (Figure 4J).
To identify the specific mutation that drives this meta- bolic phenotype (i.e. increased OXPHOS capacity in Evi1/MF9 cells), we measured OXPHOS levels and per- formed glucose/glutamine starvation experiments on cells infected with Evi1-targeting shRNA (shEvi1) or cells treat- ed with a DOT1L inhibitor (EPZ004777). We found that
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shEvi1 and EPZ004777 suppressed OXPHOS in Evi1/MF9 cells (Online Supplementary Figure S4A). When cells trans- fected with shEvi1 or treated with EPZ004777 were cul- tured in the presence of different low concentrations of glucose or glutamine, shEvi1 cells survived for a signifi- cantly longer time in lower concentrations of glutamine (Online Supplementary Figure S4B). This suggests that EVI1 is important for glutamine dependency. We performed quantitative PCR analysis of Glut-1, Idh1, and Idh2, and found that expression of Idh2 was lower in shEvi1 and in Evi1/MF9 cells treated with EPZ004777 (Online Supplementary Figure S4C). To investigate whether altered
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Figure 2. EVI1 increases oxidative phosphorylation (OXPHOS) and glycolysis dur- ing development of MLL-AF9 leukemia. (A) The basal oxy- gen consumption rate (OCR) and maximal respiratory capacity in Evi1-TG normal progenitor cells were signifi- cantly lower than those in wild-type (WT) progenitor cells. (B-D) After the 2nd plat- ing, the OCR of Evi1-TG pro- genitor cells infected with MLL-AF9 increased gradually when compared with that of WT cells. (E) After the 2nd plat- ing, the maximal respiratory
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increased gradually when compared with that of WT cells. (F) The basal and capac- ity OCR of EVI1/MF9 bone marrow cells were significant- ly higher than those of WT/MF9 cells. (G) The basal ECAR of Evi1/MF9 cells increased slightly after the 3rd plating. (H) There was no sig- nificant difference between ECAR of ex vivo leukemia cells from WT/MF9 and from Evi1/MF9 mice. All samples were collected at least in duplicate.
Evi1/MF9
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haematologica | 2020; 105(8)