Probing the ligand binding specificity of FNBP4 WW domains and interaction with FH1 domain of FMN1

Formins are a group of actin-binding proteins that mediate nascent actin filament polymerization, filament elongation, and barbed end capping function, thereby regulating different cellular and developmental processes. Developmental processes like vertebrate gastrulation, neural growth cone dynamics, and limb development require formins to function in a regulated manner. Formin binding proteins like Rho GTPase regulates activation of auto-inhibited conformation of diaphanous formins. Unlike other diaphanous formins, Formin1 (FMN1), a non-diaphanous formin, is not regulated by Rho GTPase. FMN1 acts as an antagonist of the BMP signaling pathway during limb development. Several previous reports demonstrated that WW domain-containing proteins can interact with poly-proline-rich amino acid stretches of formins and play a crucial role in developmental processes. WW domain-containing FNBP4 protein plays an essential role in limb development. It has been hypothesized that the interaction between FNBP4 and FMN1 can further attribute to the role in limb development through the BMP signaling pathway. In this study, we have elucidated the binding kinetics of FNBP4 and FMN1 using surface plasmon resonance and enzyme-linked immunosorbent assays. Our findings confirm that the FNBP4 exhibits interaction with the poly-proline-rich formin homology 1 (FH1) domain of FMN1. Furthermore, only the first WW1 domains is involved in the interaction between the two domains. Thus, this study sheds light on the binding potentialities of WW domains of FNBP4 and their possible contribution to the regulation of FMN1 function.