Fig. 1: Expression modulation of the histone H3K4 demethylase KDM5B/JARID1B.

a Anti-KDM5B immunostaining of a melanoma patient sample (left) compared to a benign human nevus (right). Highly positive nuclei are dark red, medium positive nuclei are light red, low expressing nuclei are blue. Isotype controls are shown in the upper right corners. Depicted are representative images of different melanoma or nevi samples (n = 5 each). b Kaplan–Meier survival curves of cutaneous melanoma patients were calculated from the TCGA data set based on cut-point optimization for KDM5B expression (high expression, red, vs. low, green, TCGA browser tool UCSC Xena and GraphPad Prism). Sample sizes are indicated in the patient at risk table (# of risk). Significance was tested by Long-rank (Mantel–Cox) test. c Quantitation of KDM5B mRNA induction after 24 h of doxycycline (Dox) treatment as assessed by qPCR. Shown is one representative example (mean, n = 2). d Anti-KDM5B nuclear immunostaining of WM3734^Tet3G-KDM5B cells after Dox-titration at the indicated concentrations for 24 h (left, representative images; right, quantitation shown as normalized frequency distribution of nuclear staining intensity). Shown is one representative out of three clones. e Flow cytometric detection of endogenous KDM5B protein levels after treatment with Cpd1 for 72 h. Mean ± SD (n = 4); two-sided t-test. f Anti-KDM5B nuclear immunostaining of three different melanoma cell lines (WM3734, WM88, MelJuso) after 72 h of 10 µM Cpd1 treatment (left, representative pictures; right, quantitation shown as normalized frequency distribution of nuclear staining intensity). g Time course of KDM5B protein levels after treatment of WM3734 cells with Cpd1 (10 µM) plus cycloheximide (CHX, 50 µg/ml, n = 2). h Ubiquitin protein conjugates were immunoprecipitated in WM3734 cells after Cpd1 treatment for 72 h. The total ubiquitinated cellular protein content was detected by a pan-ubiquitin-HRP antibody (upper row). The fraction of ubiquitinated KDM5B protein was detected by a KDM5B-specific antibody (middle row). Neg4 was used as structure homolog compound control (n = 2). To improve visualization of the KDM5B bands resulting from immunoprecipitation, the contrast was enhanced for both treatments equally. Source data are provided as a Source Data file.