Identification of a missense point mutation of SHIP1. (A) Schematic of the isoleucine to threonine amino acid substitution in the 5′-phosphatase domain of SHIP1 and the corresponding conserved hydrophobic residue of other inositol 5′-phosphatases. (B) 5′-Phosphatase assay showing the conversion of PtdIns(3,4,5)P₃ substrate to PtdIns(3,4)P₂ product by immunoprecipitated recombinant wild-type or mutant SHIP1. The HA vector sample is included as a transfection control, and results are representative of 2 individual experiments. The HA-immunoblot of lysates and parallel HA-immunoprecipitates, on which the 5′-phosphatase assays were performed, demonstrates similar loading of SHIP1 protein. (C) Western blot for SHIP1 expression in resting SHIP1^+/+, SHIP1^−/−, and SHIP1^el20/el20 BMMΦs. Western blot is representative of 3 independent samples for each genotype. (D) Quantitative RT-PCR for SHIP1 mRNA expression in whole bone marrow (BM), thymus (THY), sorted lineage⁻c-Kit⁺ (LK) BM cells and resting BMMΦs. All values have been normalized to expression of HPRT. Bar graphs represent the mean ± SD of 3 samples for each genotype, with the exception of LK cells, which were obtained from sorts of 3 mice for each genotype. **P < .01, 2-tailed Mann-Whitney test. ***P < .001, 2-tailed Mann-Whitney test. ND indicates not detected.