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Fig. 4 | BMC Cancer

Fig. 4

From: Methionine regulates self-renewal, pluripotency, and cell death of GIC through cholesterol—rRNA axis

Fig. 4

A Microarray results of top 10 master, maintenance, and marker genes changed by methionine depletion. Averages of the interested gene mRNA expression from the expression array are depicted using Prism7. B Validation with qPCR of master, maintenance, and marker genes of GICs. Methionine depletion down-regulated OLIG2, SOX2, SOX4, FOXM1, and PROM1 mRNA gene expression in MZGC4. C FCM (left panel) and immunoblot analysis (right panel). Methionine depletion decreased markers of GICs CD133 and SOX2. Left panel: CD133 expression in cultured medium with (green, pink) / without (black) methionine in MZGC7. Methionine depletion for 5 days (Green) and 7 days (black) reduced CD133 expression to the same level of isotype control (pink). Orange overlay showed CD133 positive cells as positive control. Right panel: Methionine depletion for 5 days reduced SOX2 expression in MZGC7 and 8. Each experiment was performed in triplicate. (Full-length blots/gels are presented in Supplementary Figure S10A). D IGV showing gene locus of PROM1 with DMRs of MZGC1, MZGC2, and MZGC3 along with gene map from Reference Sequence (RefSeq), CpGi, binding site of insulator, transcriptional factors, and histone markers from ChIP-seq database. DMRs by methionine depletion were localized in almost the same locus, where CpGi and gene body occur, in all cell lines. Close-up of the promoter region shows DMRs are localized around CpGi and shores (right). FOXM1 binds to intron of PROM1 gene body, whose histone mark is H3K27ac. Hypermethylated (pink) and hypomethylated (green) in the methionine depleted compared to control states are indicated. Hg19 used as reference genome

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