SCF-mediated c-Kit internalization and downstream signaling. (A) SCF-c-Kit internalization after SCF binding was measured at indicated time points. BMMCs generated from 3 different wild-type and Kit^787F mice were compared. One representative experiment from 3 is shown (*P = .009; **P = .001; ***P = 4 × 10⁻⁴; ****P = 2 × 10⁻⁴). (B) SCF-c-Kit internalization was induced after preincubation of cells with Dynasore (100μM) or DMSO (control) for 30 minutes at 37°C and subsequent loading with SCF (50 ng/mL). αSCF staining (white histograms, MFI indicated) and staining of isotype-matched antibodies of irrelevant specificity (gray histograms) were measured at indicated time points by flow cytometry. One representative experiment from 3 is shown. (C) BMMCs from wild-type mice were preincubated with Dynasore (100μM) or DMSO (control) for 30 minutes at 37°C and stimulated for the indicated times with SCF (100 ng/mL) or PMA and ionomycin. Cell lysates were immunoblotted with antibodies specific for (phospho-)Erk1/2, (phospho-)c-Kit, and c-Kit. Immunoblotting with Erk1/2-specific antibodies served as loading control. One representative experiment from 2 is shown.