Nuclear accumulation of Smad complexes occurs only after the midblastula transition in Xenopus.
Activin and the Nodal-related proteins induce mesendodermal tissues during Xenopus development. These signals act through specific receptors to cause the phosphorylation, at their carboxyl termini, of Smad2 and Smad3. The phosphorylated Smad proteins form heteromeric complexes with Smad4 and translocate into the nucleus to activate the transcription, after the midblastula transition, of target genes such as Xbra and goosecoid (gsc). In this paper we use bimolecular fluorescence complementation (BiFC) to study complex formation between Smad proteins both in vivo and in response to exogenous proteins. The technique has allowed us to detect Smad2-Smad4 heteromeric interactions during normal Xenopus development and Smad2 and Smad4 homo- and heteromers in isolated Xenopus blastomeres. Smad2-Smad2 and Smad2-Smad4 complexes accumulate rapidly in the nuclei of responding cells following Activin treatment, whereas Smad4 homomeric complexes remain cytoplasmic. When cells divide, Smad2-Smad4 complexes associate with chromatin, even in the absence of ligand. Our observation that Smad2-Smad4 complexes accumulate in the nucleus only after the midblastula transition, irrespective of the stage at which cells were treated with Activin, may shed light on the mechanisms of developmental timing.
Saka
Yasushi
Y
Wellcome Trust/Cancer Research UK Gurdon Institute and Department of Zoology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.
Hagemann
Anja I
AI
Piepenburg
Olaf
O
Smith
James C
JC
D028341:Activins / Q000494:pharmacology; D000818:Animals; D036703:Blastula; D002467:Cell Nucleus / Q000378:metabolism*; D004530:Egg Yolk / Q000502:physiology; D004625:Embryo, Nonmammalian / Q000502:physiology; D005260:Female; D016297:Mutagenesis, Site-Directed; D010063:Ovum / Q000502:physiology; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D051785:Smad Proteins / Q000235:genetics* / Q000378:metabolism*; D051899:Smad2 Protein / Q000235:genetics / Q000378:metabolism; D051900:Smad3 Protein / Q000235:genetics / Q000378:metabolism; D051901:Smad4 Protein / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D014981:Xenopus / Q000196:embryology*; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism
D012333:RNA, Messenger; D051785:Smad Proteins; D051899:Smad2 Protein; C497785:Smad2 protein, Xenopus; D051900:Smad3 Protein; D051901:Smad4 Protein; D029867:Xenopus Proteins; D028341:Activins