Fig. 6

Identification of direct targets of SOX11 and SOX4 by ChIP-seq profiling. a Western blot analysis of induced expression of SOX11-FLAG and SOX4-FLAG in keratinocytes. Western blotting was done on isolated GFP+ cells cultured for 24 h with or without Dox. The representative blots shown are from three independent experiments. b ChIP-qPCR verifies binding of genomic region containing known SOX4/11 binding sites in the Tead2 and Tubb3 genes to FLAG-tagged SOX11 or SOX4 protein. Primers were designed to amplify the known binding sites (positive) or neighboring regions without potential SOX consensus elements (negative). Data are the mean ± SD. n = 4 biological replicates. *p < 0.05 (Student’s one-tailed t-test). c Overlay of SOX11 and SOX4 ChIP-seq peaks identified from ChIP-seq experiment where lysates from two biological replicates of keratinocytes deficient of both Sox11 and Sox4 induced to express SOX11-FLAG or SOX4-FLAG for 24 h and the amount of specified genomic regions bound by SOX11-FLAG or SOX4-FLAG were quantified. d Representative ChIP-seq profiles of indicated genes with genomic region binding to SOX11 and SOX4. e Annotation of SOX11 and SOX4 genome-wide binding sites. f Homer motif analysis for SOX11 and SOX4 ChIP-seq. (Left) Matches to known motifs; and (Right) de novo motif enrichment with best motif matches indicated. FRA1 and FOSL2 are FOS-related members of AP-1 transcription factor complex. g SOX4, AP-1, and TEAD motif enrichment around SOX11 and SOX4 binding sites. h Ratios of SOX11 or SOX4 ChIP-seq peaks with or without SOX4 and/or AP-1 consensus binding motifs. i Genomic regions containing SOX4 and SOX11 ChIP-seq peak from Tead2, Fscn1, Fblim1, Pxdn, and Marcksl1 were cloned and placed in either forward (F) or reversed (R) orientation upstream of the luciferase reporter gene. Luciferase activity was measured and Firefly luciferase activity was normalized over Renilla luciferase activity. Graph shows normalized luciferase activity relative to vector control. n = 5 independent experiments. Data are mean ± SD. *p < 0.05, **p < 0.01 (Student’s one-tailed t-test). Source data for panels a, b, h, and j are provided as a Source Data file