Cdc42 is important for early B cell development in the bone marrow. (A) Genetic approach used to ablate Cdc42 specifically in B cells. (B and C) Bone marrow from WT and Cdc42 KO mice was analyzed by flow cytometry. The gating strategies are shown on the left. B cell progenitors were divided into 3 populations on the basis of CD43 and B220 expression levels (B220⁺CD43⁺, B220⁺CD43⁻, and mature recirculating cells [B220hiCD43⁻]) as shown in B. They were further subdivided on the basis of CD24, BP-1, IgM, and IgD expression levels into populations A(B220⁺CD43⁺CD24⁻BP-1⁻), B(B220⁺CD43⁺CD24⁺BP-1⁻), C(B220⁺CD43⁺CD24⁺BP-1⁺), D(B220⁺CD43⁻IgM⁻IgD⁻), E(B220⁺CD43⁻IgM⁺IgD^−/Int), and nonrecirculating (IgD^hiB220⁺CD43⁻IgM^+/IntIgD⁺) as shown in C. Quantifications are shown in the right-hand column and indicate percentage of cells in the indicated gates. Data were pooled from at least 4 independent experiments with at least 2 mice in each group. (D) B cell progenitors (A–E) were sorted from the bone marrow and protein expression of Cdc42 (first row) and actin (second row) were analyzed by Western blot. Densitometric analysis was used to quantify the signal intensity of Cdc42 normalized to actin and is shown in the chart below. (E) In vivo labeling of bone marrow progenitors. Intravenously injected anti CD45.2-PE was used to distinguish cells in BM sinusoid (PE⁺) from cells in BM parenchyma (PE⁻). The first two columns show a representative example of one WT mouse (blue) and one Cdc42 KO mouse (red). Populations D, E, and E’ are shown. Quantification charts are shown in the two right-hand columns; data were pooled from three independent experiments with five mice in each group. *, P < 0.05; **, P < 0.01; ****, P < 0.0001.