Figure 4. Knockdown of Foxp1 recapitulates miR-34a-induced B-lineage abnormalities.

A. Schematic diagram showing the MGP-based construct used to express the Foxp1 siRNA.

B. Left hand panel shows Foxp1 RNA, measured by RT-qPCR, in 70Z/3 cells infected with either empty vector (Vector) or MGP-Foxp1si (Foxp1si). Right hand panel shows immunoblot analysis of Foxp1 in these cells. The numbers represent the fold expression of Foxp1 at the protein level compared to vector control.

C. GFP expression in the bone marrow of mice (vector and Foxp1si) 2 months after transplant, measured by flow cytometry.

D. Foxp1 expression in the bone marrow of mice (vector and Foxp1si), as measured by RT-qPCR. Individual dots represent Foxp1 RNA amounts in individual mice, while the bars show the mean amount for a representative experiment (n=4 for each group; T-test, p=0.0026). Experiments were repeated three times, with similar trends in all three experiments.

E. Analysis of pro-B cells. Left hand panels show representative histograms of the GFP⁺IgM⁻ negative compartment in control (vector) and Foxp1si mice. The right hand panel shows pro-B cells as a percentage of total GFP+ cells for a representative experiment, with individual mice represented by individual dots and the bars indicating the mean (n=4 per group; T-test, p=0.0004). Three independent experiments showed similar trends.

F. Analysis of pre-B cells. Left hand panels show representative histograms of the GFP+ IgM− compartment in control and Foxp1si mice, below which the percentage of cells in each of the four quadrants is shown. The right hand panel shows pre-B cells as a percentage of total GFP+ cells for a representative experiment, with individual mice represented by individual dots and the bars indicating the mean (n=4 per group; T-test, p=0.005). Three independent experiments showed similar trends.