Fig. 5: Suppression of SOX10 by pharmacologic activation of WNT signaling is not transcriptional, but mediated by the proteasome.

a Representative western blots of the indicated proteins in the MAPK inhibitor resistant human melanoma cell culture M12, at different timepoints of CHIR99021 (6 μM) treatment (timepoints are shown directly in the figure). GAPDH was used as loading control (n = 3). b RNA levels in MAPK inhibitor sensitive (M98) human melanoma cell culture (ΔCt to GAPDH expression) after 4, 8, and 12 h of CHIR99021 (6 µM) treatment are shown compared with DMSO (ctrl) (n = 3). c Representative western blot of the indicated proteins in the MAPK inhibitor sensitive human melanoma cell culture (M98). A representative western blot for the indicated proteins of M98 human melanoma cell culture stably expressing shSOX10 or shMITF constructs cultured in presence or absence of CHIR99021 for 24 h, 6 μM. Nssh represents the scrambled shRNA used as control construct (n = 3). d Representative western blot of the indicated proteins in presence of CHIR99021 (6 μM) and/or MG-132 (20 μM) (in single or double treatments) for 16 h. GAPDH was used as a loading control (n = 3). e Heatmap demonstrating the gene expression of SOX10 and Wnt target genes of M98 and M00 (MAPK inhibitor sensitive cell cultures) and M11 (resistant to MAPK inhibitor) in either presence or absence of CHIR99021. Data on the heat map represent log2 of sequencing reads assigned to genes. Statistical significance was determined by unpaired, two-tailed Student’s t test in b. *P < 0.05, **P < 0.01, ***P < 0.001. In each panel, n indicates the number of independent experiments performed.