Rcor1 mutants exhibit a hypersensitive CSF2 signaling pathway. (A-B) Rcor1 and Gfi1b occupy the promoters of indicated target genes measured by (A) Rcor1 ChIP and (B) Gfi1b ChIP analysis in the MEL cell line. Binding is represented as fold enrichment relative to a negative region from the β-actin gene intron. For each gene, a site 2 to 8 kb away from the positive binding site served as the internal NC. Results from 3 experiments are shown (mean ± SD). ‡ indicates comparisons to β -actin: ‡P < .05; ‡‡P < .01; ‡‡‡P < .001. *Indicates comparison with internal NC: *P < .05; **P < .01; ***P < .001. (C) Rcor1 ChIP and (D) Gfi1b ChIP analysis in primary control R2 fetal liver cells (FLCs) and mutant total FLCs. Binding is represented as fold enrichment relative to a negative region from the β-actin gene intron. The mean of 2 independent experiments is shown. (E) Representative flow cytometry analysis of Csf2rb expression level in E13.5 control fetal liver (n = 10) and mutant fetal liver (n = 5). PE-conjugated anti-Csf2rb or PE-conjugated immunoglobulin G1 (isotype control) were used. (F) Western blot analysis showing that purified TER119^– mutant cells have higher levels of p-Stat5, but similar levels of total Stat5 and Jak2 protein, following treatment with CSF2. This experiment was repeated once more with the same result. (G) Colony-forming assay results showing that the Jak2 inhibitor TG101348 reduces CFU-GM colonies generated from mutant fetal liver R2 cells. Results from 4 independent control or mutant fetal livers treated with TG101348 or dimethylsulfoxide (DMSO) are shown (mean ± SD). ***P < .001.