Fig 1: SOX2 inhibits DNMT1-dependent maintenance of DNA methylation during replication.a In vitro methylation assay to measure DNMT1 methyltransferase activity using hemi-methylated probes containing WT or Sc PF motifs in the presence or absence of the corresponding PF. Relative DNMT1 activity is represented as scintillation counts in WT probes, corrected for the amount of recovered DNA probes and compared relative to the Sc probe. Results are shown as mean + SEM of n = 3 biologically independent replicates. SOX2, ETS1 and OTX2 significantly reduce DNMT1 activity. p values: ETS1 p = 0.0018, OTX2 p = 0.0238, SOX2 p = 0.026 (two-tailed unpaired t test; *p < 0.05, **p < 0.01). b In vitro methylation assay using the probe containing the OCT4SOX2 motif in the presence of OCT4, SOX2 or SOX2 + OCT4 proteins. Results are shown as mean + SEM of n = 3 biologically independent replicates. p values: SOX2 + OCT4 p = 0.018, SOX2 p = 0.003, (two-tailed unpaired t test, *p < 0.05, **p < 0.01). c, d In vitro replication assay to assess the effect of SOX2 and OCT4 binding on the maintenance of DNA methylation during replication. Two probes containing the OCT4SOX2 motif were used: FR1 (c) and FR2 (d). The indicated concentrations represent those of active hOCT4 and hSOX2 recombinant proteins that were used. Methylation levels following replication are measured based on the integration of radioactively labelled methyl group during replication and compared to “no_protein” control. Results are presented as mean + SD of n = 5 (hSOX2 samples) or n = 3 biologically independent replicates (hOCT4 and hOCT4 + hSOX2 samples) and analysed as radioactive signal in the presence of the protein relative to the signal in the absence of protein. p values: FR1_600nM_SOX2 p = 0.0092, FR1_600nM_OCT4 + SOX2 p = 0.0068, FR2_450nM_SOX2 p = 0.0071, FR2_600nM_SOX2 p = 0.0021, FR2_450nM_OCT4 + SOX2 p = 0.0026, FR2_600nM_OCT4 + SOX2 p = 0.0047 (two-tailed unpaired t test, **p < 0.01). For all panels, source data are provided as a Source data file.