Dissection of inhibitory Smad proteins: both N- and C-terminal domains are necessary for full activities of Xenopus Smad6 and Smad7

• Published in: Mechanisms of development Vol. 100; no. 2; pp. 251 - 262
• Main Authors: Nakayama, T, Berg, L K, Christian, J L
• Format: Journal Article
• Language: English
• Published: Ireland 01.02.2001
• Subjects: Animals; Blotting, Western; Bone Morphogenetic Proteins - metabolism; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - genetics; DNA-Binding Proteins - physiology; Gene Deletion; Models, Genetic; Mutation; Phenotype; Plasmids - metabolism; Protein Structure, Tertiary; Reverse Transcriptase Polymerase Chain Reaction; RNA - metabolism; Signal Transduction; Smad6 Protein; Smad7 Protein; Spinal Dysraphism - metabolism; Structure-Activity Relationship; Time Factors; Trans-Activators - chemistry; Trans-Activators - genetics; Trans-Activators - physiology; Xenopus; Xenopus Proteins
• ISSN: 0925-4773
• DOI: 10.1016/s0925-4773(00)00533-5

Smad6 and Smad7 comprise a subclass of vertebrate Smads that antagonize, rather than transduce, TGF-beta family signaling. These Anti-Smads can block BMP signaling, as evidenced by their ability to induce a secondary dorsal axis when misexpressed ventrally in Xenopus embryos. Smad7 inhibits additional TGF-beta related pathways, and causes spina bifida when misexpressed dorsally. We have performed structure-function analyses to identify domains of Anti-Smads that are responsible for their shared and unique activities. We find that the C-terminal domain of Smad7 displays strong axis inducing activity but cannot induce spina bifida. The isolated N-terminal domain of Smad7 is inactive but restores the ability of the C-terminus to cause spina bifida when the two are co-expressed. By contrast, the N- and C-terminal domains of Smad6 have weak axis inducing activity when expressed individually, but show full activity when co-expressed. Chimeric analysis demonstrates that the C-terminal domain of Smad7, but not Smad6, can induce spina bifida when fused to the N-terminal domain of either Smad6 or Smad7. Thus, although the C-terminal domain is the primary determinant of the intrinsic activity of Xenopus Anti-Smads, the N-terminal domain is essential for full activity, is interchangeable between Smad6 and 7, and can function in trans.