Fig. 7: FOXK1 O-GlcNAcylation regulates its transcriptional function on chromatin.

a Chromatin occupancy of Flag-tagged FOXK1, FOXK1^7A and FOXK1^11A on all human promoters in IMR90 cells. b Bar-plot representing the repartition of endogenous FOXK1 and exogenous (3 Flag-tagged) FOXK1, FOXK1^7A and FOXK1^11A on the genome of IMR90 cells assessed by ChIP-seq. c, d Venn diagram depicting the overlap in chromatin occupancy between FOXK1, FOXK1^7A, and FOXK1^11A at promoters and at distal regions in IMR90 cells. e Top: Pulldown assay using HeLa nuclear extracts incubated with recombinant GST-FOXK1, which was either pre-modified by in vitro O-GlcNAcylation or incubated with OGT without UDP-GlcNAc (Representative of three independent experiments). Bottom: Quantification of BAP1 band signal intensity in pulldown conditions with or without FOXK1 O-GlcNAcylation in vitro (n = 4 independent experiments). f Differential recruitment of BAP1 in IMR90 cells overexpressing FOXK1, FOXK1^7A or FOXK1^11A. Regions were identified by comparing the list of BAP1 peaks from the FOXK1 expressing condition to those from the FOXK1^11A condition. Technical replicates were merged for visualization. Boxplot representing BAP1 spike-in normalized read counts. Endogenous FOXK1 was depleted by siRNA in conditions expressing FOXK1 and O-GlcNAc mutants. Two technical replicates were merged. g GO analysis performed on promoters (249) differentially enriched for BAP1 between FOXK1 and FOXK1^11A. h Boxplot representing H3K4me1 and H2AK119ub spike-in normalized reads in IMR90 cells expressing FOXK1, FOXK1^7A or FOXK1^11A on regions with differential BAP1 recruitment. Two technical replicates were merged. Statistics: Data are represented as mean ± SEM (e). One-way ANOVA with two-sided tests followed by Tukey’s multiple comparisons test for adjustment (f, h). Two-tailed Mann–Whitney U test was used to compare the two groups P = 0.0286 (e). GO enrichment analysis was performed using Enrichr. Statistical significance was assessed using Fisher’s exact test, and odds ratios (OR) were computed (g). Box plots display the normalized read counts for each condition. The center line represents the median. The box extends from the 25th to the 75th percentile (interquartile range, IQR). The whiskers extend to the most extreme non-outlier values within 1.5 × IQR. Outliers are not shown (f, h). *P  <  0.05. Source data are provided as a Source Data file.