Fig. 6: SOX2 and GLI1 co-regulate ST3GAL1 gene transcription. | Nature Communications

Fig. 6: SOX2 and GLI1 co-regulate ST3GAL1 gene transcription.

From: ST3GAL1 is a target of the SOX2-GLI1 transcriptional complex and promotes melanoma metastasis through AXL

Fig. 6

a Schematic representation of ST3GAL1 regulatory regions (RRs). b, c ChIP-qPCR of SOX2 and GLI1 occupancy of the ST3GAL1 RRs showing that both TFs bind to ST3GAL1 enhancer II in SSM2c (b) and A375 M6 (c) cells. The y-axis represents relative promoter enrichment, normalized on input material. IgG was set to 1. ACTIN promoter was used as negative control. Data are represented as mean ± s.e.m. P value was calculated by two-tailed unpaired Student’s t test (n = 3 biological independent experiments); ns not significant. d Consensus SOX2 and GLI1 binding sites identified in ST3GAL1 enhancer II. e, f Quantification of dual-luciferase reporter assay in SSM2c melanoma cells showing that silencing of GLI1 or SOX2 is able to counteract the transactivation induced by SOX2 (e) or GLI1 (f) on the enhancer II of ST3GAL1. Relative luciferase activities were firefly/Renilla ratios, with level induced by control equated to 1. Data are represented as mean ± s.e.m. P value was calculated using ANOVA and Tukey’s test (n = 4 in e, n = 5 in f). g Western blot of SOX2, GLI1, and ST3GAL1 in A375 M6 and SSM2c cells transduced as indicated. h Quantification of dual-luciferase reporter assay in SSM2c cells showing that mutagenesis of GLI1-BS and SOX2-BS-2 prevents SOX2 and GLI1 from transactivating ST3GAL1 enhancer II. Relative luciferase activities were firefly/Renilla ratios, with the level induced by the control equated to 1. Data are represented as mean ± s.e.m. by two-tailed unpaired Student’s t test (n = 4). i Matrigel invasion assay in A375 M6 cells transduced as indicated. Data are represented as mean ± s.d. using two-sided Kruskal–Wallis and Dwass-Steel-Critchlow-Fligner method (n = 3 biological independent experiments). j Western blot of SOX2, GLI1, ST3GAL1, and EMT markers in A375 M6 cells transduced as indicated. k Representative images of i. Blots in g and j are representative of n = 3 biological independent experiments. HSP90 was used as loading control. Source data are provided as Source Data file.

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