Variant of A431 cells isolated by ricin A-conjugated monoclonal antibody directed to EGF receptor: phosphorylation of EGF receptor and phosphatidylinositol.
A monoclonal antibody specific for human EGF receptors was cross-linked to subunit A of toxic ricin. Using this conjugate, we isolated a variant of A431 cells, designated C1-B7, with approximately 40 times less EGF binding capacity. Unlike the parental cells, the C1-B7 variant was resistant to EGF-induced suppression of cell growth. The EGF receptors retained in this variant were of high-affinity type and susceptible to EGF-induced autophosphorylation. Membrane prepared from C1-B7 cells was highly phosphorylated in the presence of 2 microM [gamma-32P]-ATP, primarily on the lipid components shown as phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate. This same level of lipid phosphorylation was observed on A431 membrane only in the presence of higher ATP concentrations. After addition of EGF to A431 membrane, phosphatidylinositol phosphorylation was significantly decreased with a concomitant increase in EGF-dependent protein phosphorylation. Thus, the EGF-dependent receptor-mediated protein phosphorylation precedes phosphatidylinositol phosphorylation. These observations support the idea that the growth inhibitory effect of EGF on A431 cells is caused by high ATP consumption due to the EGF-induced protein phosphorylation and reduction of phosphatidylinositol turnover.
D000911:Antibodies, Monoclonal; D002294:Carcinoma, Squamous Cell; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002469:Cell Separation; D004261:DNA Replication / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000378:metabolism* / Q000494:pharmacology; D066246:ErbB Receptors; D014644:Genetic Variation; D006801:Humans; D007700:Kinetics; D010716:Phosphatidylinositols / Q000378:metabolism*; D010766:Phosphorylation; D011956:Receptors, Cell Surface / Q000378:metabolism*; D012276:Ricin / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D010716:Phosphatidylinositols; D011956:Receptors, Cell Surface; D004815:Epidermal Growth Factor; D012276:Ricin; D066246:ErbB Receptors