Calmodulin differentially modulates Smad1 and Smad2 signaling

The members of the Smad protein family are intracellular mediators of transforming growth factor beta (TGF-beta) signaling. Smad1 transduces bone morphogenetic protein signals, inducing formation of ventral mesoderm in Xenopus embryos, whereas Smad2 transduces activin/TGF-beta signals, generating dorsal mesoderm. Calmodulin directly binds to-many Smads and was shown to downregulate Smad2 activity in a cell culture system (Zimmerman, C. M., Kariapper, M. S. T., and Mathews, L. S. (1997) J. Biol. Chen. 273, 677-680). Here, we extend those data and demonstrate that calmodulin alters Smad signaling in living embryos, increasing Smad1 activity while inhibiting Smad2 function. To characterize this regulation, we undertook a structure-function analysis and found that calmodulin binds to two distinct and conserved;regions in both Smad1 and Smad2. Receptor tyrosine kinase signaling also modifies Smad activity (Kretzschmar, M., Doody, J., and Massague, J. (1997) Nature 389, 618-622; Kretzschmar, M Doody, J., Timokhina, I., and Massague, J. (1999) Genes Dev. 13, 804-816; de Caestecker, M. P., Parks, W. T., Frank, C. J., Castagnino, P., Bottaro, D. P., Roberts, A. B., and Lechleider, R. J. (1998) Genes Dev. 12, 1587-1592). We show that calmodulin binding to Smads inhibits subsequent Erk2-dependent phosphorylation of Smads and vice versa. These observations suggest the presence of a cross-talk between three major signaling cascades as follows: Ca2+/calmodulin, receptor tyrosine kinase, and TGF-beta pathways.