Abstract

Addition of leupeptin, methylamine and the antitubulin agent nocodazole did not affect the initial rate of association of 125I-labelled epidermal growth factor (125I-EGF) to Swiss mouse 3T3 fibroblast cells in vitro, but continued incubation with these drugs (up to 24 h) led to an increase in cell-associated radioactivity in a time- and dose-dependent fashion. Combinations of these drugs caused additive increments in cell-associated and internalized radioactivity. Throughout the incubation period, 81-89% of the cell-associated 125I-EGF was internalized. Upon incubation of 125I-EGF with 3T3 cells in the presence or absence of the three inhibitors of degradation for periods of up to 24 h, and after removal of the surface-bound material, the internalized 125I-EGF was extracted and 42-53% was found to biochemically intact (by acid precipitation) and 56-65% was antigenically similar to native EGF (using double antibody immunoprecipitation in an EGF radioimmunoassay). The extracted internalized 125I-EGF was capable of binding to fresh 3T3 cells. Despite causing a similar increase in intact internalized 125I-EGF, leupeptin did not interfere with and nocodazole alone or in combination with leupeptin markedly enhanced EGF-stimulated DNA synthesis, whereas methylamine inhibited mitogenesis. These data indicate a dissociation between EGF degradation and DNA synthesis, and are not consistent with the hypothesis that intracellular degradation of EGF is necessary for its mitogenic effects.

View details for Web of Science ID A1982PS24800014

View details for PubMedID 6982826