Decreased binding of epidermal growth factor to BALB/c 3T3 mutant cells defective in glycoprotein synthesis

Abstract

THE surface of animal cells contains a variety of complex macromolecules, some of which modulate cell growth, adhesion and motility. A number of peptide hormones and growth factors influence cellular proliferation and macromolecular synthesis with binding to surface proteins of responsive cells as the first phase of their action^1–4. The structure and identity of these receptor molecules is a topic of widespread interest. Epidermal growth factor (EGF) is a low molecular weight peptide (MW 6,000) capable of stimulating DNA synthesis and proliferation in epidermal cells⁵, human fibroblasts⁶, and mouse embryo fibroblasts (3T3)⁷. It has also been reported to stimulate the synthesis of a major cell surface glycoprotein (CSP)⁸ and hyaluronic acid⁹ in 3T3 cells and human fibroblasts maintained in low serum. These effects occur at extremely low concentrations of EGF, in the ng ml⁻¹ range, and the first step in this process is the specific binding to surface receptors, followed by rapid internalisation and degradation¹⁰. Pouyssegur and Pastan¹¹ have isolated a nontransformed mutant (AD6) of BALB/c 3T3 cells that has a dramatic decrease in cell surface carbohydrates due to a specific, but partial block in the acetylation of N-acetylglucosamine-6-phosphate. This early defect in the biosynthesis of amino sugars leads to incomplete glycosylation of glycoproteins and to decreased exposure of glycoproteins on the cell surface¹². Culturing the cells in the presence of N-acetylglucosamine (GlcNAc) bypasses the enzymatic block and results in a restoration of altered surface components¹³. We report here that the mutant AD6 cells show a dramatic decrease in EGF binding, whereas their ability to bind insulin is normal. Our data are consistent with the notion that the receptor for EGF is a glycoprotein.