Activation of MAP-Kinase and PI-3Kinase/Akt in erythroid progenitors by stromal cells expressing either the wt-SCF²²⁰ (MA) orSl^17H-SCF²²⁰ (MA) isoform of SCF. Factor-starved erythroid progenitors were cocultured with mitomycin C-treated stromal cells expressing either the wt or theSl^17H MA SCF for various time points. Subsequently, at various times, cell lysates were collected and subjected to Western blot analysis with a rabbit anti-phospho Akt antibody and a rabbit anti-phospho MAP-Kinase antibody and the enhanced chemiluminescence detection system. The phosphorylated form of Akt (upper panel) and MAP-Kinase (Erk-1 and Erk-2) (middle panel) are indicated. In both the upper and middle panels, lane 1 corresponds to unstimulated starved erythroid progenitor cells and lanes 2, 3, and 4 correspond to stromal cells expressing the wt-SCF²²⁰ (MA) form of SCF cocultured with erythroid progenitor cells for 10, 60, and 120 minutes, respectively. Lanes 5, 6, and 7 correspond to stromal cells expressing the Sl^17H-SCF²²⁰ (MA) form of SCF cocultured with erythroid progenitor cells for 10, 60, and 120 minutes, respectively. The bottom panel demonstrates the loading control for total protein in each lane. These experiments were performed at least 3 times with 2 different clones of each genotype.