Epidermal growth factor stimulates the synthesis of cell-attachment proteins in the human breast cancer cell line PMC42

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Abstract
The human breast cancer cell line PMC42 responds to the addition of epidermal growth factor (EGF) by proliferation and increased frequency of attachment of cell-organoid structures to the culture vessel. Antibodies to fibronectin and laminin reacted strongly, by immunoperoxidase, with the membranes of cells from organoids that became adherent following addition of EGF. This reaction was weak with membranes of cells of non-adherent organoids in cultures containing EGF and was negative with membranes of cells of free-floating organoids from cultures without EGF. An increase in biosynthetic labelling with 35S-methionine was found in cell lysates and supernatants of PMC42 cells cultured in the presence of EGF compared with control cells grown without EGF. One-dimensional SDS-polyacrylamide gel electrophoresis and 2-dimensional NEPHGE-PAGE of labelled cell proteins showed increased synthesis of several cellular proteins and the appearance in EGF-treated cells of 2 proteins which were not detected in cells from control cultures lacking EGF. Immunoprecipitation experiments using antibodies to fibronectin and laminin with lysates of 35S-methionine labelled PMC42 cells cultured with EGF showed strong immunoprecipitation at Mr 200 and 400 with anti-laminin, and at Mr 200 and 96 with antifibronectin. These immunoprecipitates were blocked specifically by purified laminin or fibronectin, respectively. No immunoprecipitates were detected with these antibodies in lysates from cells grown without EGF. EGF thus stimulates increased adherence of cultured PMC42 cell-organoid structures together with increased membrane expression of the cell-adhesive proteins laminin and fibronectin. These effects may play a role in normal development and neoplastic behaviour of breast epithelia.