Fig 1: IDH mutations activate CEBPα-VDR-RXRα axis through 2HG production. (A) RT-qPCR showing gene expressions of VDR and RARA in HL60 IDH1WT (clone 2: ○; clone 7: ☐) versus HL60 IDH1R132H (clone 5: ◇; clone 11: △). (B) Western blot showing protein levels of VDR, RXRα and RARα in HL60 IDH1WT (clones 2 and 7) versus HL60 IDH1R132H (clones 5, 11 and 3) (left) and in HL60 IDH2WT versus HL60 IDH2R172K (right). (C) RT-qPCR showing gene expressions of VDR and RARA in 2HG-treated HL60 (200 µM, 1 week). (D) Western blot showing protein levels of VDR RXRα and RARα in 2HG-treated HL60 IDH1WT (clones 2 and 7) (200 µM, 1 week). (E) RT-qPCR showing gene expression of and VDR and RARA in HL60 IDH1WT (clone 2: ○, clone 7: ☐) and IDH1R132H (clone 5: ◇, clone 11: △) after CEBPA-KD by shRNA (F) Western blot showing protein levels of CEBPα, VDR, RARα and VDR in HL60 IDH1WT (clones 2 and 7) and IDH1R132H (clone 5 and 11) after CEBPA-KD by shRNA. (G) Western blot showing protein levels of VDR in 2HG-treated HL60 IDH1WT (clone 2 and 7) with CEBPA-KD by shRNA (200 µM, 1 week). The uncropped western blot figures are presented in Figure S6. * p < 0.05; ** p < 0.01.
Fig 2: Vitamin D receptor-related gene signatures are enriched in transcriptomes of IDH mutant AML cells. (A) Gene signature of pharmacological substances were compared with gene signatures associated with IDH1MUT HL60 cells, IDH1WT HL60 cells+2HG and IDHMUT patient cells from TCGA and GSE14468 transcriptomes using Genomatix software. VENN diagram represents common pharmacological substances associated with these four gene signatures. (B) VENN diagram of common genes from ATRA- and VD-responsive genes (Pharmacological substances, Genomatix) enriched in IDHMUT-AML and HL60 IDH1WT+2HG. (C) Gene set enrichment analysis (GSEA) of late VD/VDR activation pathway signature in HL60 IDH1R132H, HL60 IDH1WT treated with 2HG and in IDHMUT patients from GSE14468. (D) Correlation between late VD/VDR activation pathway gene signature and up-regulated genes in IDH1MUT signatures (left, GSE14468; right, TCGA) in scRNA-seq of AML cells from the PDX IDH1R132H. (E) GSEA of late VD/VDR activation pathway gene signature in IDHMUT patients relapsing after IDH inhibitor treatment compared to those patients before treatment (GSE153349).
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