Recruitment of Actin-Binding Proteins on the Membrane Interface: Effects of Cholesterol on Protein/PIP2 Interactions

Abstract

Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) only represents 1% of membrane phospholipids but is a key ligand in a myriad of important physiological events such as membrane-cytoskeleton interactions. PI(4,5)P2 activates various actin-binding proteins that regulate actin microfilament assembly which in turns plays an essential role in dynamic changes in the cytoskeleton. Understanding PI(4,5)P2's ability to bind numerous actin-binding proteins that contain structurally divergent PI(4,5)P2 binding sites will provide invaluable insights linking molecular-level specificity to cytoskeletal assembly and dynamics. However, the underlying atomistic mechanisms by which PI(4,5)P2 interacts with several actin-binding proteins has not been well investigated. Cholesterol's role in mediating peptide and lipid interactions is also not fully understood. Previously, we have shown both computationally and experimentally that lipid composition, specifically PI(4,5)P2 concentration and cholesterol presence regulates the ability for PI(4,5)P2 to interact with an actin nucleating protein, Mdia2 (a formin protein). In this current study, we employed several all-atom molecular dynamics (MD) simulations on three systems, namely, peptides derived from the formin protein mDia2, neural wiskott-aldrich syndrome (N-Wasp), an actin nucleating protein, and actin filament severing protein, gelsolin. The peptides were adhered on lipid bilayers containing varying levels of PI(4,5)P2 and cholesterol concentration. Remarkably, our results reveal electrostatic interactions between the peptides and lipid bilayers are uniquely specific to structure of the peptide, PI(4,5)P2 concentration, and cholesterol presence. Analysis of the electrostatic interactions PI(4,5)P2 and formin and gelsolin peptide indicate that the presence of cholesterol, increases PI(4,5)P2 recruitment. Dynamic light scattering experiments performed on the peptides and bilayers were found to be in agreement of our MD simulations. These results suggest a new understanding of the underlying mechanisms of peptide binding to the membrane in a physiologically relevant bilayer environment mediated by PI(4,5)P2 and cholesterol.