Mechanochemical coupling of formin-induced actin interaction at the level of single molecular complex

Formins promote actin assembly and are involved in force-dependent cytoskeletal remodeling. However, how force alters the formin functions still needs to be investigated. Here, using atomic force microscopy and biomembrane force probe, we investigated how mechanical force affects formin-mediated actin interactions at the level of single molecular complexes. The biophysical parameters of G-actin/G-actin (GG) or G-actin/F-actin (GF) interactions were measured under force loading in the absence or presence of two C-terminal fragments of the mouse formin mDia1: mDia1Ct that contains formin homology 2 domain (FH2) and diaphanous autoregulatory domain (DAD) and mDia1Ct-Delta DAD that contains only FH2. Under force-free conditions, neither association nor dissociation kinetics of GG and GF interactions were significantly affected by mDia1Ct or mDia1Ct-Delta DAD. Under tensile forces (0-7 pN), the average lifetimes of these bonds were prolonged and molecular complexes were stiffened in the presence of mDia1Ct, indicating mDia1Ct association kinetically stabilizes and mechanically strengthens bonds of the dimer and at the end of the F-actin under force. Interestingly, mDia1Ct-Delta DAD prolonged the lifetime of GF but not GG bond under force, suggesting the DAD domain is critical for mDia1Ct to strengthen GG interaction. These data unravel the mechanochemical coupling in formin-induced actin assembly and provide evidence to understand the initiation of formin-mediated actin elongation and nucleation.