Fig 2: Proposed model for LKB1-mediated m6A modification in KRAS-mutated lung cancer progression.Loss of LKB1-induced DNA hypermethylation, which prevents CTCF binding on the ALKBH5 gene promoter, maintains ALKBH5 expression, and further represses global RNA methylation. Oncogenic SMAD7, SOX2, and MYC are crucial targets of m6A meditated by LKB1 deficiency, and are involved in KRAS mutation lung cancer progression.

Fig 3: Suppressor-CTCF is required for ALKBH5 downregulation by LKB1.A–C Representative images and quantification of CTCF and ALKBH5 protein expression by WB and qRT-PCR in A549 cells with LKB1 overexpression and/or CTCF knockdown for 48 h. Error bars, SD (n = 4 for WB; n = 5 for qRT-PCR), *P < 0.05 vs. Si-CN or OE-CN (one-way ANOVA with Bonferroni multiple comparison post hoc test). D Luciferase reporter assay of A549 cells transfected with pGL3-basic constructs containing serial LKBH5 promoters or deletion of CTCF peak fragment. n = 8/group, mean ± SD. * P < 0.05 by one-way ANOVA followed by Tukey’s test. E Reduction of ALKBH5 Luc:-1168 bp wild-type (WT) construct activities by treatment of 5-aza or LKB1 overexpression in A549 cells, but not for ALKBH5 Luc:-1168 bp deletion (Del). Data as mean ± SD (n = 5), *P < 0.05 (Student’s t-test). F ChIP-qPCR showing that the ALKBH5-CTCF peak region was occupied by the suppressor of CTCF and activators of histone modulators in A549 cells. G–J ChIP-qPCR analyses of 5-aza treated or LKB1 overexpressed A549 cells. n = 5/group, mean ± SD. *P < 0.05 vs. IgG in F, vs. A549-DMSO in G–J by one-way ANOVA followed by Tukey’s test.

Fig 4: m6A level significantly decreased during osteogenic differentiation of hASCs. a Representative image of m6A level quantification assay. b Quantitative analyses of m6A level. c Expression of m6A regulators during the osteogenic differentiation of hASCs by RNA-seq (uhASCs vs dhASCs-7 day). d qRT-PCR analyses of the expression of METTL3, METTL14, FTO, and ALKBH5. e Western blot analysis of METTL3, METTL14, FTO, and ALKBH5 (uhASCs vs dhASCs-7 day). *p < .05, **p < .01, ***p < .001. hASCs, human adipose-derived stem cells; qRT-PCR, quantitative real-time polymerase chain reaction; uhASCs, undifferentiated hASCs; dhASCs, osteogenically differentiated hASCs

Fig 5: HSP90 inhibitor-induced temporal changes in protein levels. Western blot images and the quantification data showing the alterations in expression levels of the HSP70, DNAJB4, METTL3, FTO, ALKBH5, and YTHDF3 in M14 cells at different time points following treatment with 100 nM ganetespib (a), AT13387 (b), or 17-DMAG (c). Shown are the ratios of expression of the indicated proteins over ß-actin, and further normalized to the ratios obtained for the control cells without HSP90 inhibitor treatment. The data represent the mean ± S.D. of results from three independent experiments. The p-values referred to the comparison between 0 h and 24 h treatment and were calculated using unpaired, two-tailed Student’s t-test: #p > 0.05; *, 0.01 = p < 0.05; **, 0.001 = p < 0.01; ***, p < 0.001. Source data are provided as a Source Data file