DNA methyltransferase 1/miR-342-3p/Forkhead box M1 signaling axis promotes self-renewal in cervical cancer stem-like cells in vitro and nude mice models

Figure 2 DNA methyltransferase 1 and forkhead box M1 as the direct targets of miR-342-3p in HeLa-derived cervical cancer stem cell-like cells. A: Heatmaps depict the relative expression levels of all 29 tumor microRNAs (miRNAs) in HeLa cells and their responding cervical cancer stem cell-like cells (CCSLCs). The color scale represents the expression levels of miRNA as upregulation (red), downregulation (green), or middle expression (light color) across all samples. Lower volcano plot analysis showed differentially expressed (P < 0.005, student’s t-test, log₂ fold change > 2) miRNA in HeLa cells and their responding CCSLCs; B: Visual representation displaying the expression levels of all 92 genes investigated in this study through heatmap analysis. Scale: Relative expression levels were normalized for internal control. Lower volcano plot showing the differentially expressed mRNAs (P < 0.005, student’s t-test, log₂ fold change > 2) in HeLa-derived CCSLCs treated with miRNA (miR)-negative control or miR-342 (n = 3); C: MiR-342-3p promoter methylation in HeLa cells and their CCSLCs; D: The relative luciferase activity in HeLa-derived CCSLCs was determined after the DNA methyltransferase 1 (DNMT1) 3’-untranslated regions (3’-UTRs) or mutant (MUT) plasmids were cotransfected with miR-342-3p mimics or inhibitors or negative control; E: The relative luciferase activity in HeLa-derived CCSLCs was determined after the forkhead box M1 (FOXM1) 3’ UTR or mutant plasmids were cotransfected with miR-342-3p mimics or inhibitors or the negative control; F: DNMT1 mRNA level in HeLa cells and its corresponding CCSLCs; G: MiR-342-3p level in HeLa cells and its corresponding CCSLCs; H: FoxM1 protein amounts in HeLa cells and its corresponding CCSLCs, with α-tubulin as a loading control; I: Schematic representation of the pathomechanism by which DNMT1/miR-342-3p/FOXM1 axis promotes self-renewal-related stemness in CCSLCs. ^aP < 0.05 vs HeLa-derived cervical cancer stem cell-like cells cotransfected with DNMT1 3’-UTR with miR-negative; ^bP < 0.05 vs HeLa-derived cervical cancer stem cell-like cells cotransfected of mutant plasmid in the presence or absence of miR-342 mimics (n = 3). WT: Wildtype; M: Methylated; U: Unmethylated.