MUTATIONAL ANALYSIS OF A TRANSFORMING-GROWTH-FACTOR-BETA RECEPTOR-BINDING SITE

Transforming growth factor-beta s (TGF-beta 1,beta 2,beta 3) are impor tant regulators of cell growth and differentiation which share approxi mately 70% identical amino acids. Using LS513 colorectal cells, which are growth inhibited by TGF-beta 1 (ED50 of 100 pM), but are refractor y to TGF-beta 2 (ED50 of 50,000 to 100,000 pM), we have determined tha t amino acids 92-98 of TGF-beta specify growth inhibition. The chimeri c protein TGF-beta 1/beta 2(92-98), in which amino acids 92-98 of TGF- beta 1 were exchanged for the corresponding amino acids of TGF-beta 2, was indistinguishable from TGF-beta 2 at inhibiting growth of LS513 c ells. In contrast, both TGF-beta 1/beta 2(92-95) and TGF-beta 1/beta 2 (94-98) inhibited the growth of LS513 cells with an ED50 of approximat ely 1000 pM. TGF-beta 1/beta 2(95-98), in which amino acids 95-98 of T GF-beta 1 have been replaced with the corresponding amino acids of TGF -beta 2, had full activity and was indistinguishable from TGF-beta 1. Receptor cross-linking experiments demonstrated that binding of the ch imeras to the type I and type Il receptors of LS513 cells was consiste nt with their biological activity. TGF-beta 1/beta 2(92-98), TGF-beta 1/beta 2(92-95) and TGF-beta 1/beta 2(94-98) were each similar to TGF- beta 2 in that they failed to bind to the soluble Type II receptor in a solid-phase assay. These results demonstrate that amino acids 92-98 are involved in the interaction between TGF-R and its signaling recept ors and they show that modest changes within this region can substanti ally alter biological response.