Expression and tyrosine phosphorylation of 32D infectants.

The 32D cells were infected with the indicated c-kitconstructs, selected, sorted, and cloned. These clones were then measured for receptor levels by flow cytometry using biotinylated-SLF, followed by streptavidin-PE. Levels of c-Kit receptor on KitWT, KitYF719, KitYF728, and KitYF719/YF728 cells were approximately equivalent as measured by the mean fluorescence intensity (MFI). Uninfected 32D cells are represented by the shaded histogram. (B): c-Kit receptors from KitWT, KitYF719, KitYF728, and 32D cells were either not stimulated (−) or stimulated (+) with SLF for 2.5 minutes at 37°C. Receptors were then precipitated and resolved by 7.5% SDS-PAGE, followed by transfer to nitrocellulose and blotting with antiphosphotyrosine antibodies (middle panel). Blots were then stripped and reprobed with anti-p85 antibodies (upper panel). Equal levels of Kit were precipitated in each sample as evidenced by stripping and reprobing with anti–c-Kit antibodies (bottom panel).