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. Author manuscript; available in PMC: 2008 Jun 24.
Published in final edited form as: Development. 2007 Oct 24;134(23):4209–4218. doi: 10.1242/dev.010645

Fig. 2. Tagged forms of Smad2 and Smad4 retain their biological activities.

Fig. 2

(A) Illustration of the principle of bimolecular fluorescence. In unstimulated cells, in which Smad2 is not phosphorylated, Smad2 and Smad4 do not interact, and the N-terminal and C-terminal portions of VENUS are not able to complement. Phosphorylation of Smad2 results in a conformational change and allows interaction with Smad4 and fluorescence complementation. (B) Tagged Smad constructs retain their biological activity. Xenopus embryos received no injections (embs, caps), or were injected at the one cell stage with RNA encoding tagged forms of Smad2 or Smad4 proteins (100 pg for VC155-Smad2/VNm9-Smad4 BiFC tags and 80 pg for HA tags), alone or in combination, or with 5 pg RNA encoding Activin. Animal caps were dissected at the midblastula stage and cultured to the equivalent of early gastrula stage 10.5. Note that when expressed individually, tagged forms of Smads do not induce expression of Xbra and goosecoid, but coexpression of Smad2 and Smad4 (100 pg RNA of each) activates both genes at levels similar to those induced by Activin.