Gene expression profiling and a gain-of-function screen identify potential regulators of HSPC colonization. (a) GSEA enrichment plots for the chemokine gene set in embryonic and adult endothelial cells and HSCs. In both plots, genes enriched in endothelial cells are plotted to the left and genes enriched in HSCs are plotted to the right (P = 0.000 for both comparisons). (b) Overlap of the leading-edge chemokine genes from the embryonic and adult GSEA. (c–g) Leading edge genes were induced by heat shock at 36 and 48 hpf, followed by fixation at 72 hpf and WISH using a mix of runx1/c-myb probes to mark HSCs and HSPCs. Representative animals injected with heat shock inducible plasmids encoding GFP (c), cxcr1 (d), and cxcl8 (f) are shown. Bars, 100 µm. Bar plots (e and g) show blinded scoring data for runx1/c-myb staining in the CHT of CXCR1 (e) and CXCL8 (g) groups compared with GFP control (Wilcoxon rank sum test, P = 0.033, n = 43 for GFP control, and n = 20 for CXCR1; P = 0.0028; n = 15 for GFP control and n = 26 for CXCL8). Both experiments were repeated twice with similar results. Representative experiments are shown. To account for clutch-to-clutch variability in staining, all experimental groups were compared only to controls from the same clutch. (h and i) WISH for cxcr1 expression in a WT 48 hpf embryo. The images are representative of two separate clutches. Bars, 700 µm. (j) Expression of cxcl8 and cxcr1 mRNA in endothelial cells freshly sorted from 72 hpf kdrl:mCherry embryos. (k and l) Sorted endothelial cells were treated with EET or DMSO (control) for 30 min before assessment of cxcl8 (k) and cxcr1 (l) expression by qRT-PCR. Experiments in j–l were repeated twice, with similar results.