Figure 4

Tol scaffold containing the BMP2 knuckle peptide, KIPKASSVPTELSAISTLYL, activates Smad dependent signalling both when added exogenously to adherent cells and when immobilised on a surface. (a) Cells were cultured in multiwell plates, transfected with a SMAD-reporter construct and then treated with recombinant BMP2, exogenous Tol-BMP-ST or Tol-ST. Levels of luciferase were then assessed after eight hours. Both recombinant BMP2 (31 nM) and Tol-BMP-ST (112 nM) induced significant levels of luciferase activity compared to Tol alone. (b) Cells already transfected with the SMAD-reporter construct were plated onto gold coverslips displaying the scaffold proteins at the concentrations outlined in Table 1 or TEG alone. Luciferase activity was measured 12 hours later and contrasted with the same cells grown on tissue culture plastic and treated with soluble recombinant BMP-2 (100 ng/ml). Significant levels of SMAD signalling were only observed on the Tol-BMP-ST surface. Data representative of three independent experiments; results are mean ± SEM; ** P < 0.01. Significance was determined by ANOVA with Tukey post test.