Figure 2

Phosphorylation and immobilization of proteins associated with the integrin-cytoskeleton-complex. Cytoskeletally anchored α_Vβ₆ was immunoprecipitated after TGFβ₁ stimulation (10 nM for 10 minutes) followed by Western analysis with antibodies against tyrosine-phosphorylated proteins (A) or Western blotting after biotinylation of all proteins and streptavidin detection (B). Presence of TGFβ₁ (C), α_V and β₆ integrin (D) in the co-precipitates is also demonstrated. TGFβ-receptor-I and II (TGFβRI and TGFβRII) are expressed at nearly equal levels in all cell lines as demonstrated by western blotting from whole cell extracts (E). In part the cells were preincubated with α_V- and β₆-antibodies (1:100 each for 30 min) or with a TGFβ antibody (15 μg/ml for 30 min).