Fig. 4
From: The WNT target SP5 negatively regulates WNT transcriptional programs in human pluripotent stem cells
SP5 regulates expression of developmental regulators. a Altered gene expression in WNT-treated SP5 mutant cells. A heatmap of gene expression illustrates differences between WT and SP5 mutant (dZF1 and 2) hESCs. Genes are in order of the clusters identified in Fig. 1c; expression is given as percent WT maximum RPKM (0, white; 100, blue; ≥100, red). Refer to Supplementary Data 3 for complete gene list. b Representative upregulated genes. The average change in percent maximum RPKM of dZF 1 and dZF 2 vs. wild-type illustrates upregulation of many developmental regulators (−1, indigo; 0, white; 1.0, red). Genes in red contain a SP5 peak. *SP5 peaks near FOXA2 and SOX17 were called at a lower confidence (p < 10−4 vs. p < 10−5) and were not included in the list of 662 SP5 peaks. c SP5 peaks are enriched among genes with the most significant fold changes upon WNT treatment. For each gene set (A through D), the table provides the p-value for the least significantly changed gene (as described by Trapnell et al.52, 53). Refer to Supplementary Data 4 for gene lists. The graph indicates the fraction of genes with SP5 peaks among gene sets A through D. d Genes with SP5 peaks are associated with early embryonic development. Gene ontology (GO) analysis reveals that genes with SP5 peaks of gene set C are highly enriched for genes associated with “primitive streak formation”. e Validation of upregulated genes. WT and dZF mutant hESCs were treated with 1 nM Wnt3a for the indicated times and RNA was isolated and analyzed by qPCR for expression of SP5. Genome browser tracks illustrate the increase of SP5 binding near the transcriptional start site. Additional examples are provided in Supplementary Fig. 5. Arrows indicates direction of transcription. RQ relative quantity. (Error bars are SEM for four technical replicates; Student’s t test: *p < 0.05; **p < 0.01; ****p < 0.0001). f Extended Wnt3a treatment of hPSC. WT and SP5 mutant (dZF2) cells were treated with Wnt3a for the indicated times and RNA was analyzed by qPCR for SP5 expression
