The mechanisms by which the binding of peptide mitogens to their receptors results in initiation of DNA synthesis are poorly understood. Lectins such as Concanavalin A (Con A) and wheat germ agglutinin (WGA) may be useful probes for studying mitogen action in human fibroblasts. Studies done in this laboratory suggest that these carbohydrate-binding proteins inhibit the mitogenic effects of growth factors such as epidermal growth factor (EGF) and somatomedin C (Sm-C) with little or no effect on mitogen binding. This proposal will investigate mechanisms by which lectins may induce post-receptor refractoriness to mitogen action in fibroblasts. First, the effect on WGA on four different mitogen-stimulated events, Na+ influx, glucose transport, amino acid transport and DNA synthesis will be compared to better define the biochemical basis for its inhibitory effects. Second, studies will investigate the paradoxical effects of Con A, which enhances fibroblast amino acid uptake as do other mitogens, but inhibits rather than stimulates DNA synthesis. Third, the effects of WGA, Con A and other lectins on binding and internalization of I125-EGF and I125-Sm-C will be examined to determine the relationship, if any, between these effects and inhibition of mitogen action. In light of evidence that clustering of mitogen receptors is required to initiate a biological effect, and that lectins can inhibit receptor mobility, it is hypothesized that lectins block mitogen action by preventing aggregation of mitogen-receptor complexes. In the last series of experiments, the effect of lectins on the clustering of EGF receptors tagged with an EGF-ferritin conjugate will be studied by electron microscopy, and evidence for a role of the microtubule-microfilament network in mediating the inhibitory effects of lectins will be sought. These studies should contribute to our understanding of normal cell growth regulation by allowing a distinction between cellular events which can be induced by mitogen binding alone, and those which require a post-binding event, such as receptor clustering, to be intiated. Furthermore, the ability of certain lectins to inhibit human fibroblast growth stimulation without cell toxicity, in a manner which is reversible by addition of the appropriate saccharide, may have useful applications in the study of rapid or uncontrolled human cell replication.
| Kaplowitz, P B; Haar, J L (1988) Antimitogenic actions of lectins in cultured human fibroblasts. J Cell Physiol 136:13-22 |
| Kaplowitz, P B (1987) Glucocorticoids enhance somatomedin-C binding and stimulation of amino acid uptake in human fibroblasts. J Clin Endocrinol Metab 64:563-71 |
