Combined loss of cIAPs and XIAP does not impair myeloid cell differentiation but impairs survival of cells in macrophage colonies. (A) Representative fluorescence-activated cell sorter plots with percentages of surface marker staining to identify myeloid progenitors in the bone marrow of WT, c1^fl/flc2^−/−X^−/−, and c1^LCc2^−/−X^−/− mice. Cells were gated for Sca1^–c-kit⁺ and further gated for FcγRcII/III and CD34 (CMP, common myeloid progenitor; GMP, granulocyte macrophage progenitor; MEP, megakaryocyte erythrocyte progenitor). (B) Spleen cells from newborn WT, c1^fl/flc2^−/−X^−/−, and c1^LCc2^−/−X^−/− mice were plated in agar with either GM-CSF, Multi-CSF (GM-, G-, M-CSF), or stem cell factor (SCF) + IL-3 + Epo, and the numbers and morphologies of the colonies (Eo, eosinophil; G, granulocytes; GM, granulocyte-macrophage; M, macrophage; Meg, megakaryocyte) determined after 7 days. N = 3 biological replicates. (C) BMDMs and peritoneal macrophages were treated with the indicated concentrations of the Smac mimetic Comp. A and assayed 24 hours for cell death by flow cytometry. (D) BMDMs from mice of the indicated genotypes were either left untreated (-) or treated with Comp. A (500 nM, 24 hours; +). The percentages of dead cells were determined by flow cytometric analysis after staining with PI. N = 3 experiments with BMDMs from individual mice. (E) BMDMs from mice of the indicated genotypes were left untreated (open bars) or treated for 24 hours with either 500 nM Comp. A, a Smac mimetic that inhibits all of cIAP1, cIAP2, and XIAP, or with 500 nM of a Smac mimetic that only inhibits cIAP1 and cIAP2. The percentages of dead cells were determined by staining with PI followed by flow cytometric analysis. N = 3 experiments with BMDMs from individual mice of the genotypes indicated. Mean ± SEM are shown. **P < .01; *P < .05.