Disruption of transforming growth factor beta signaling by a mutation that prevents transphosphorylation within the receptor complex

T beta R-II (transforming growth factor beta [TGF-beta] type II receptor) is a transmembrane serine/threonine kinase that acts as the primary TGF-beta receptor. Ligand binding to T beta R-II leads to the recruitment and phosphorylation of T beta R-I, a distantly related transmembrane kinase that acts as a downstream signaling component. T beta R-I phosphorylation by T beta R-II is shown here to be essential for signaling. A mutant T beta R-II that binds ligand but lacks signaling activity was identified. This mutant was identified by screening with a TGF-beta-inducible vector a series of mink lung epithelial cell clones that have normal TGF-beta binding activity but have lost antiproliferative and transcriptional responses to TGF-beta. When transiently cotransfected with T beta R-II, one of these cell lines, S-21, recovered TGF-beta responsiveness. cDNA cloning and sequencing of T beta R-II from S-21 cells revealed a point mutation that changes proline 525 to leucine in kinase subdomain XI. A recombinant receptor containing this mutation, T beta R-II(P525L), is similar to wild-type T beta R-II in its abilities to bind ligand, support ligand binding to T beta R-I, and form a complex with T beta R-I in vivo. T beta R-II(P525L) has autophosphorylating activity in vitro and in vivo; however, unlike the wild-type receptor, it fails to phosphorylate an associated T beta R-I. These results suggest that T beta R-II(P525L) is a catalytically active receptor that cannot recognize T beta R-I as a substrate. The close link between T beta R-I transphosphorylation and signaling activity argues that transphosphorylation is essential for signal propagation via T beta R-I.

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