Mode of Ezrin-Membrane Interaction as a Function of PIP 2 Binding and Pseudophosphorylation

Abstract

Ezrin, a protein of the ezrin, radixin, moesin (ERM) family, provides a regulated linkage between the plasma membrane and the cytoskeleton. The hallmark of this linkage is the activation of ezrin by phosphatidylinositol-4,5-bisphosphate (PIP2) binding and a threonine phosphorylation at position 567. To analyze the influence of these activating factors on the organization of ezrin on lipid membranes and the proposed concomitant oligomer-monomer transition, we made use of supported lipid bilayers in conjunction with atomic force microscopy and fluorescence microscopy. Bilayers doped with either PIP2 as the natural receptor lipid of ezrin or a Ni-nitrilotriacetic acid-equipped lipid to bind the proteins via their His6-tags to the lipid membrane were used to bind two different ezrin variants: ezrin wild-type and ezrin T567D mimicking the phosphorylated state. Using a combination of reflectometric interference spectroscopy, atomic force microscopy, and Förster resonance energy transfer experiments, we show that only the ezrin T567D mutant, upon binding to PIP2-containing bilayers, undergoes a remarkable conformational change, which we attribute to an opening of the conformation resulting in monomeric protein on the lipid bilayer.

Pentachloropseudilin Impairs Angiogenesis by Disrupting the Actin Cytoskeleton, Integrin Trafficking and the Cell Cycle

“…Ezrin is a protein of the ezrin, radixin, moesin (ERM) family involved in adhesion, cell migration, and angiogenesis, and acts as a linker between the plasma membrane and the cortical actin cytoskeleton . The hallmark of this linkage is the activation of ezrin by phosphatidylinositol‐4,5‐bisphosphate (PIP2) binding . It is known that Myo1s exist as monomers containing a three‐part domain structure similar to that found in most myosin proteins: an N‐terminal motor domain that binds to actin and hydrolyses ATP, a neck (lever‐arm) region that consists of several calmodulin‐binding (CaM‐binding) IQ (isoleucine–glutamine) motifs, and a C‐terminal tail homology 1 (TH1) domain with a pleckstrin homology (PH) domain capable of binding to lipid membranes such as PIP2 .…”

“…[25] The hallmark of this linkagei st he activation of ezrin by phosphatidylinositol-4,5-bisphosphate (PIP2)b inding. [26] It is known that Myo1se xist as monomers containing at hree-part domain structure similart ot hat found in most myosin proteins:a nN -terminal motor domain that binds to actin and hy- drolyses ATP, an eck (lever-arm) region that consists of several calmodulin-binding (CaM-binding) IQ (isoleucine-glutamine) motifs, and aC -terminal tail homology 1( TH1) domain with a pleckstrin homology (PH) domain capable of binding to lipid membranes such as PIP2. [27] In this sense, we hypothesize that allosterici nhibition by PClP promotes small conformational changes in Myo1s and that this impairs the interaction of a Myo1 domain capable of binding to PIP2, thus compromising the activation of ezrin and inhibiting blood vessel formation.…”

Receptor‐Free Signaling at Curved Cellular Membranes

“…[5,62,63] Within the framework of curvature-sensing proteins, lipid dependence has been demonstrated for the BAR domain proteins Pacsin 1, Pacsin 2, and FBP17 (to PS or PIP and PS [59] ). [5,62,63] Within the framework of curvature-sensing proteins, lipid dependence has been demonstrated for the BAR domain proteins Pacsin 1, Pacsin 2, and FBP17 (to PS or PIP and PS [59] ).…”

Maspin and Ezrina - Biomarker Molecules in Colorectal Cancer Correlative immunohistochemical study

“…EBP50, CD44, ICAM-2) via the N-terminus end. In the active state, N-terminus end binds NHERF (Na+/H+ exchanger regulatory factor) [11].…”