DEVELOPMENT OF EXPRESSION CLONING SYSTEM FOR ONCOGENE CDNAS

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An expression cloning strategy was devised to isolate the keratinocyte growth factor (KGF) receptor cDNA by creation of an autocrine transforming loop. NIH/3T3 fibroblasts, which secrete this epithelial cell-specific mitogen, were transfected with a keratinocyte expression cDNA library. Among several transformed foci identified, one demonstrated the acquisition of specific high affinity KGF binding sites. The pattern of binding competition by related fibroblast growth factors (FGFs) indicated that this receptor had high affinity for acidic FGF (aFGF) as well as KGF. The rescued 4/2-kb cDNA was shown to encode a predicted membranespanning tyrosine kinase related to but distinct from the basic FGF (bFGF) receptor. This expression cloning approach may be generally applicable to the isolation of genes that constitute limiting steps in mitogenic signalling pathways.