Fig. 3. Stimulation of Fn assembly is mediated by interaction of surface tTG with the gelatin-binding domain of Fn. (A,B) tTG-stimulated increase in [¹²⁵I]Fn incorporation into the deoxycholate-insoluble fraction is inhibited by the 42 kDa Fn fragment and anti-tTG antibody. (A) Deoxycholate-insoluble [¹²⁵I]Fn in the matrix of Swiss 3T3 transfectants grown for 24 hours with 50 nM [¹²⁵I]Fn was analyzed by reducing SDS-PAGE and autoradiography. (B) Quantitation of the [¹²⁵I]Fn incorporation into the deoxycholate-insoluble fraction. ¹²⁵I-labeled bands corresponding to Fn monomer and polymer were cut out of the gels and counted. The results are representative of three independent experiments (mean±s.e.m.). (C) tTG-mediated increase in Fn fibril formation by tTG[2] and tTG_C277S[2] transfectants is blocked by the 42 kDa fragment and anti-tTG antibody. Cells grown for 24 hours with 50 nM exogenous Fn were stained with anti-Fn antibody. Bar, 50 µm. (A-C) Where indicated, cells were grown in the presence of 20 µg/ml anti-tTG antibody, blocking anti-5ß1 integrin mAb BMA5, control nonimmune IgG or 2 µM 42 kDa or 110 kDa fragments of Fn.