Gfi1b regulates Gfi1 and the expression of surface molecules important for niche organization. (A) MxCre tg Gfi1b^fl/fl Gfi1^GFP/wt, wt, and Gfi1b^fl/fl Gfi1^GFP/wt mice were injected with pIpC, and 30 days after the first injection (equivalent to 21 days after the last injection) mice were killed and examined for expression of GFP, which follows the activity of the Gfi1 promoter. Loss of Gfi1b leads to an enhanced activity of the Gfi1 promoter. (B) Real-time polymerase chain reaction analysis of Gfi1 gene expression in HSCs from mice with the indicated genotypes (n = 3). (C) Example of the clustering of Gfi1b effector genes enriched in a specific geneset (Adhesion molecules) generated using the GSEA software (http://www.broad.mit.edu/gsea/).(D) Overview of genes differentially expressed in wt and Gfi1b-deficient HSCs. Red bars represent relatively high and blue bars low expression levels (average fold induction or repression) in Gfi1b^ko/ko HSCs compared with wt HSCs. CXCR4 and VCAM-1 were not included in the GESA defined adhesion molecule pathway but were also down-regulated on the RNA level. (E) Expression level of different surface adhesion proteins. The expression of these proteins was changed in analogy to the gene expression array results. Mean fluorescence intensities (MFIs) of the respective surface molecules in Gfi1b^ko/ko (ko) and wt HSCs (wt) are depicted. Dotted line indicates isotype controls.