title: A novel phosphatidylinositol 4,5-bisphosphate binding domain mediates plasma membrane localization of ExoU and other patatin-like phospholipases.
abstract: Bacterial toxins require localization to specific intracellular compartments following injection into host cells. In this study, we examined the membrane targeting of a broad family of bacterial proteins, the patatin-like phospholipases. The best characterized member of this family is ExoU, an effector of the Pseudomonas aeruginosa type III secretion system. Upon injection into host cells, ExoU localizes to the plasma membrane, where it uses its phospholipase A2 activity to lyse infected cells. The targeting mechanism of ExoU is poorly characterized, but it was recently found to bind to the phospholipid phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), a marker for the plasma membrane of eukaryotic cells. We confirmed that the membrane localization domain (MLD) of ExoU had a direct affinity for PI(4,5)P2, and we determined that this binding was required for ExoU localization. Previously uncharacterized ExoU homologs from Pseudomonas fluorescens and Photorhabdus asymbiotica also localized to the plasma membrane and required PI(4,5)P2 for this localization. A conserved arginine within the MLD was critical for interaction of each protein with PI(4,5)P2 and for localization. Furthermore, we determined the crystal structure of the full-length P. fluorescens ExoU and found that it was similar to that of P. aeruginosa ExoU. Each MLD contains a four-helical bundle, with the conserved arginine exposed at its cap to allow for interaction with the negatively charged PI(4,5)P2. Overall, these findings provide a structural explanation for the targeting of patatin-like phospholipases to the plasma membrane and define the MLD of ExoU as a member of a new class of PI(4,5)P2 binding domains.
authors.affiliation: From the Departments of Microbiology-Immunology.
authors.forename: Gregory H
authors.initials: GH
authors.lastname: Tyson
authors.affiliation: Biochemistry and Center for Structural Genomics of Infectious Diseases, Northwestern University, Chicago, Illinois 60611 and.
authors.forename: Andrei S
authors.initials: AS
authors.lastname: Halavaty
authors.affiliation: the Department of Chemistry, University of Illinois at Chicago, Chicago, Illinois 60607.
authors.forename: Hyunjin
authors.initials: H
authors.lastname: Kim
authors.affiliation: From the Departments of Microbiology-Immunology.
authors.forename: Brett
authors.initials: B
authors.lastname: Geissler
authors.affiliation: From the Departments of Microbiology-Immunology.
authors.forename: Mallory
authors.initials: M
authors.lastname: Agard
authors.affiliation: From the Departments of Microbiology-Immunology.
authors.forename: Karla J
authors.initials: KJ
authors.lastname: Satchell
authors.affiliation: the Department of Chemistry, University of Illinois at Chicago, Chicago, Illinois 60607.
authors.forename: Wonhwa
authors.initials: W
authors.lastname: Cho
authors.affiliation: Biochemistry and Center for Structural Genomics of Infectious Diseases, Northwestern University, Chicago, Illinois 60611 and.
authors.forename: Wayne F
authors.initials: WF
authors.lastname: Anderson
authors.affiliation: From the Departments of Microbiology-Immunology, Medicine, and ahauser@northwestern.edu.
authors.forename: Alan R
authors.initials: AR
authors.lastname: Hauser
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism*; D010740:Phospholipases / Q000378:metabolism*; D020637:Photorhabdus / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011550:Pseudomonas aeruginosa / Q000378:metabolism; D011551:Pseudomonas fluorescens / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; C111749:pseudomonas exoprotein A protein, Pseudomonas aeruginosa; D010740:Phospholipases
keywords: Bacterial Pathogenesis; Bacterial Toxin; Crystal Structure; Phosphatidylinositol Signaling; Phospholipase; Pseudomonas aeruginosa (P. aeruginosa)