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.
Cárcamo
J
J
Cell Biology and Genetics Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021.
Zentella
A
A
Massagué
J
J
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D016384:Consensus Sequence; D004261:DNA Replication / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D005353:Fibronectins / Q000096:biosynthesis; D007668:Kidney; D008168:Lung; D008322:Mammals; D008907:Mink; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010748:Phosphopeptides / Q000737:chemistry / Q000302:isolation & purification; D010766:Phosphorylation; D017395:Plasminogen Activator Inhibitor 1 / Q000096:biosynthesis; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D018125:Receptors, Transforming Growth Factor beta / Q000096:biosynthesis / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D005353:Fibronectins; D010748:Phosphopeptides; D017395:Plasminogen Activator Inhibitor 1; D018125:Receptors, Transforming Growth Factor beta; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D013755:Tetradecanoylphorbol Acetate