Characterization of the binding of multiplication-stimulating activity to a receptor for growth polypeptides in chick embryo fibroblasts.

Multiplication-stimulating activity (MSA) is the name given to a family of polypeptides with a molecular weight of approximately 10,000 that have been isolated from the culture medium of a rat liver cell line. MSA stimulates growth and DNA synthesis in chick embryo fibroblasts and has weak insulin-like metabolic activity. We have purified MSA and characterized the binding of ‘““I-labeled MSA to specific receptors in chicken fibroblasts. Binding was rapid and reversible. Competitive binding data obtained at steady state and plotted according to Scatchard were linear. A K,l of 4.2 X lo8 M-I and a binding capacity of 58,000 MSA molecules/ cell were calculated. Only unlabeled MSA or closely related polypeptides competed for the binding of MSA tracer. MSA preparations at different stages of purification inhibited binding in accordance with their relative activities as stimulators of chicken fibroblast DNA synthesis. NSILA-s (nonsuppressible insulin-like activity soluble in acid ethanol), a polypeptide purified from human plasma with chemical and biological properties similar to those of MSA, potently inhibited the binding of ‘9-labeled MSA to chicken fibroblasts. Insulin and proinsulin were 60 and 25% as potent as MSA by weight, respectively, as inhibitors of MSA binding to chicken fibroblasts. NSILA-s also effectively inhibited MSA binding to rat liver plasma membranes, whereas neither insulin nor proinsulin inhibited MSA binding to liver membranes even at high peptide concentrations. Thus, the specificity of the chicken fibroblast MSA receptor differed from that of the liver membrane MSA receptor. The specificity of the chicken fibroblast MSA receptor also differed from that of insulin receptors in chicken fibroblasts and other cells. Specifically, MSA, NSILA-s, and proinsulin were considerably more potent as inhibitors of MSA binding to MSA receptors than of insulin binding to insulin receptors. The fact that the polypeptides competing for MSA binding to chicken fibroblasts MSA, NSILA-s, insulin, and proinsulin-also promoted growth and DNA synthesis in chicken fibroblasts at comparable concentrations, further supports our contention that the MSA receptor mediates growth stimulation by these polypeptides.