Fig. 1

Protein expression and activity of KDM6A substitution and deletion variants. A Results of the ELISA-based activity assay of KDM6A WT and substitution variants are displayed as normalized absorption on the y-axis, which positively correlates with demethylated product. Calculated amount of protein is depicted on the x-axis in logarithmic scale. WT (black) and substitutions variants in TPR (blue) and IDR (grey) generate more demethylated product with rising input amounts. Substitutions affecting the JmjC domain (yellow) remain mostly at baseline levels. T726K activity (#4A) increases with input amount at 24 h (grey boxes) but not at 48 h (grey stars). Error bars represent the standard deviation between the triplicates of each individual experiment (n = 4–18, depending on variant) C Graphic display of KDM6A truncation variants. B/D Western blot showing all substitution and deletion variants 48 h after transient transfection, detected with a-eGFP (CST) and a-KDM6A (CST) antibodies, as well as WDR5 (CST) as lysate control. Variants with IDR deletions are not detectable with the a-KDM6A antibody, as it recognizes epitopes within the IDR. The ΔIDR variant was enriched via IP. Color code blue-grey-yellow according to the site of mutation (as depicted in A and C)