Tyr⁵⁶⁴ phosphorylation is critical for catalytic activity and biologic functions of SHP-1. (A) CD34⁻KSL cells from me^v/me^v mice were retrovirally transduced with empty vector or various SHP-1 mutants. Sorted GFP⁺ cells were cultured in the presence of IL-3 and SCF for 4 weeks and then measured for SHP-1 phosphatase activity. Data are mean ± SD, *P < .05 versus vector control. SHP-1 levels shown are taken from Figure 5B. (B) Stat5 phosphorylation in the cells from panel A was analyzed by flow cytometry. Stat5 levels in the cell panel were assessed by immunoblotting. (C) CD34⁻KSL cells from me^v/me^v mice were transduced with a bicistronic retroviral vector encoding wt SHP-1, DYF SHP-1, Y536F SHP-1, Y564F SHP-1, or empty vector, together with GFP. Transduced cells were sorted and cultured in the presence of IL-3 and SCF. (D) Transduced me^v/me^v CD34⁻KSL cells were adoptively transferred (without sorting) to lethally irradiated C57BL/6-Ly5.1 mice. Donor-derived (Ly5.2⁺) CD11b⁺ cells in peripheral blood were analyzed by flow cytometry.