Smad2 phosphorylation by type I receptor: contribution of arginine 462 and cysteine 463 In the C terminus of Smad2 for specificity.
Transforming growth factor-beta (TGFbeta) is a potent regulator of cell proliferation, differentiation, motility, and apoptosis. TGFbeta binds to and activates serine/threonine kinase receptors that phosphorylate Smad2 and Smad3 intracellular signal transducers at two C-terminal serine residues. Here we show that substitutions of Arg-462 and Cys-463 residues, which are in proximity of the C-terminal serine residues, inhibited TGFbeta type I receptor-dependent phosphorylation of the C-terminal Smad2 peptides and full-length GST-Smad2 proteins in vitro. In vivo, mutation of Arg-462 and Cys-463 inhibited TGFbeta1-stimulated phosphorylation of the C-terminal serine residues in Smad2. Moreover, Smad2 with mutated Arg-462 and Cys-463 was less efficient in activation of the Smad2-responsive activin-responsive element-containing luciferase reporter ARE-luc, as compared with the wild-type protein. Thus, Arg-462 and Cys-463, which are in proximity of the C-terminal serine residues, contribute to recognition and phosphorylation of the C terminus of Smad2 by type I TGFbeta receptor.
Yakymovych
Ihor
I
Ludwig Institute for Cancer Research, Box 595, SE-751 24 Uppsala, Sweden.
Heldin
Carl-Henrik
CH
Souchelnytskyi
Serhiy
S
D030201:Activin Receptors, Type I / Q000378:metabolism*; D019943:Amino Acid Substitution; D000818:Animals; D001120:Arginine; D001665:Binding Sites; D019556:COS Cells; D002522:Chlorocebus aethiops; D003545:Cysteine; D004268:DNA-Binding Proteins / Q000378:metabolism*; D051379:Mice; D009154:Mutation; D041681:NIH 3T3 Cells; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000378:metabolism*; D051899:Smad2 Protein; D013379:Substrate Specificity; D015534:Trans-Activators / Q000378:metabolism*
D004268:DNA-Binding Proteins; D018125:Receptors, Transforming Growth Factor beta; D051899:Smad2 Protein; C497783:Smad2 protein, mouse; D015534:Trans-Activators; D001120:Arginine; D017346:Protein Serine-Threonine Kinases; D030201:Activin Receptors, Type I; D000077293:Receptor, Transforming Growth Factor-beta Type I; D003545:Cysteine