title: Phosphoinositide Binding Inhibits Actin Crosslinking and Polymerization by Palladin.
abstract: Actin cytoskeleton remodeling requires the coordinated action of a large number of actin binding proteins that reorganize the actin cytoskeleton by promoting polymerization, stabilizing filaments, causing branching, or crosslinking filaments. Palladin is a key cytoskeletal actin binding protein whose normal function is to enable cell motility during development of tissues and organs of the embryo and in wound healing, but palladin is also responsible for regulating the ability of cancer cells to become invasive and metastatic. The membrane phosphoinositide phosphatidylinositol (PI) 4,5-bisphosphate [PI(4,5)P2] is a well-known precursor for intracellular signaling and a bona fide regulator of actin cytoskeleton reorganization. Our results show that two palladin domains [immunoglobulin (Ig) 3 and 34] interact with the head group of PI(4,5)P2with moderate affinity (apparent Kd=17μM). Interactions with PI(4,5)P2decrease the actin polymerizing activity of Ig domain 3 of palladin (Palld-Ig3). Furthermore, NMR titration and docking studies show that residues K38 and K51, which are present on the β-sheet C and D, form salt bridges with the head group of PI(4,5)P2. Moreover, charge neutralization at lysine 38 in the Palld-Ig3 domain severely limits the actin polymerizing and bundling activity of Palld-Ig3. Our results provide biochemical proof that PI(4,5)P2functions as a moderator of palladin activity and have also identified residues directly involved in the crosslinking activity of palladin.
authors.affiliation: Chemistry Department, Wichita State University, 1845 Fairmount Street, Wichita, KS 67260, USA. Electronic address: yadav_rahul@ku.edu.
authors.forename: Rahul
authors.initials: R
authors.lastname: Yadav
authors.affiliation: Chemistry Department, Wichita State University, 1845 Fairmount Street, Wichita, KS 67260, USA. Electronic address: rxvattepu@wichita.edu.
authors.forename: Ravi
authors.initials: R
authors.lastname: Vattepu
authors.affiliation: Chemistry Department, Wichita State University, 1845 Fairmount Street, Wichita, KS 67260, USA. Electronic address: moriah.beck@wichita.edu.
authors.forename: Moriah R
authors.initials: MR
authors.lastname: Beck
mesh_terms: D000199:Actins / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D062105:Molecular Docking Simulation; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D011485:Protein Binding; D055503:Protein Multimerization / Q000187:drug effects*
chemical_list: D000199:Actins; D003598:Cytoskeletal Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; D010750:Phosphoproteins; C485873:palladin protein, mouse
keywords: NMR; actin cytoskeleton; electrostatics; phospholipid; regulation