Fig 2: Inhibition of sphere formation and CSC marker levels in NRF2-silenced HCT116 cells. (A) HCT116 cells were transfected with the non-specific siRNA (siCtrl) or NRF2-specific siRNA (siNRF2) and grown for 3 days in sphere culture condition. Levels of NRF2, GCLC, AKR1C1, and NQO1 mRNAs were determined in the siCtrl and siNRF2 spheres. aP < 0.05 compared to the siCtrl group. (B) The siCtrl and siNRF2 HCT116 were grown in sphere culture condition, and sphere formation was assessed. Number of spheres over 70 µm diameter was counted using an image processing ToupView software. Values represent the mean ± SEM from three independent experiments. aP < 0.05 compared to the siCtrl group. (C) Levels of KLF4 and ABCG2 mRNA were assessed in the siCtrl and siNRF2 spheres. aP < 0.05 compared to the siCtrl group. All values in RT-PCR analysis represent the mean ± SEM from at least three independent experiments. NT, non-transfection group.

Fig 3: KLF4 overexpression blocks sh-OTUD1-increased NSCLC cell progression. (a) PC9 cells were transfected with pcDNA3.1 vector or vector containing KLF4. Then the expression level of KLF4 was measured by western blot assay. (b) PC9 cells were transfected with pcDNA3.1 vector or vector containing KLF4. Cell proliferation was then measured by CCK-8 assay (p < 0.01). (c) PC9 cells were transfected with pcDNA3.1 vector or vector containing KLF4. Then the migration ability of PC9 cells was measured by transwell assay (p < 0.01). (d) PC9 cells were transfected with pcDNA3.1 vector or vector containing KLF4. Then the invasion ability of PC9 cells was measured by transwell assay (p < 0.01)

Fig 4: Defective Tfcp2l1 Thr177 phosphorylation impairs the function of Tfcp2l1 in ESC differentiation and cellular reprogramming AReal-time qPCR analysis of pluripotency- and lineage-specific genes in embryoid bodies (EBs). mESCs infected with lentiviruses containing Tfcp2l1-WT (wild-type), Tfcp2l1-T177A, or Tfcp2l1-T177E constructs were used for EB formation. Lentivirus with no inserted coding sequence was used as the control. Expression levels are represented as the ratio to 0-day-old EBs of the control group.BA schematic overview of the Oct-4 locus in gcOct-4-GFP, along with the experimental design. DE, distal enhancer; PE, proximal enhancer; PP, proximal promoter.CImmunostaining of SSEA-1 protein (red) in Oct-4 + (green) germline cells in 7-day-old EBs obtained from gcOct-4-GFP ESCs infected with Tfcp2l1-WT, T177A, or T177E lentivirus (×200 magnification, scale bar = 100 µm). A merged image with higher magnification (×400 magnifications, scale bar = 100 µm) is shown in the right panel. Nuclei were counterstained with DAPI (blue).DNumber of GFP+/SSEA-1+ germ cells in the EBs on the indicated days of culture.EAn experimental overview (upper panel) and quantification (lower panel) of primed to naïve reprogramming as determined using epiblast-derived stem cells (EpiSCs) from OG2 mice, which carry the heterozygous Oct-4-GFP (?PE) transgene.FCell morphology, Oct-4-GFP expression, and AP staining (×100 magnification, scale bar = 200 µm) in iPSC colonies generated from Sox-2, Oct-4, Klf4, and c-Myc (SOKM)-containing lentivirus-infected OG2-MEFs in the absence or presence of the indicated Tfcp2l1 ORFs or shRNA lentiviruses.G, HQuantitation of AP+ iPSC colonies expressing Tfcp2l1 and its variants (G) or with Tfcp2l1 silencing (H).Data information: Values are displayed as means ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001 compared with control group; # P < 0.005, ### P < 0.001 compared with Tfcp2l1-WT. The statistical tests used are as follows: one-way (G, H) and two-way ANOVA (A, D, and E) with Bonferroni post hoc tests. Number of biological replicates is n = 3. The exact P-values and number of replicates can be found in the source data.Source data are available online for this figure.

Fig 5: The signaling pathway change of miR-7 deficiency in ALI. FVB/N6 mice (WT, n = 6) and miR-7KD mice (n = 6) were administered with i.p. 10 mg/kg LPS, respectively. After 48 h, the expression level of ERK, p-ERK, AKT, p-AKT, and p-NF-kB in lung tissue were determined with Western Blot (A) and calculated (B). (C) The expression of NF-?B was also detected by the immunohistochemical staining. Representative data of three independent experiments were shown. *p < 0.05, **p < 0.01. (D) Schematic representation of the underlying mechanism of miR-7 deficiency on ALI. miR-7 deficiency leads to upregulation of KLF4, which could successively alter the transduction of NF-?B, ERK, and AKT signaling pathway and contribute to the ameliorated pathologies of ALI.