title: Rac1 modification by an electrophilic 15-deoxy Δ(12,14)-prostaglandin J2 analog.
abstract: Vascular endothelial cells (ECs) are important for maintaining vascular homeostasis. Dysfunction of ECs contributes to cardiovascular diseases, including atherosclerosis, and can impair the healing process during vascular injury. An important mediator of EC response to stress is the GTPase Rac1. Rac1 responds to extracellular signals and is involved in cytoskeletal rearrangement, reactive oxygen species generation and cell cycle progression. Rac1 interacts with effector proteins to elicit EC spreading and formation of cell-to-cell junctions. Rac1 activity has recently been shown to be modulated by glutathiolation or S-nitrosation via an active site cysteine residue. However, it is not known whether other redox signaling compounds can modulate Rac1 activity. An important redox signaling mediator is the electrophilic lipid, 15-deoxy-Δ(12,14)-prostaglandin J2 (15d-PGJ2). This compound is a downstream product of cyclooxygenase and forms covalent adducts with specific cysteine residues, and induces cellular signaling in a pleiotropic manner. In this study, we demonstrate that a biotin-tagged analog of 15d-PGJ2 (bt-15d-PGJ2) forms an adduct with Rac1 in vitro at the C157 residue, and an additional adduct was detected on the tryptic peptide associated with C178. Rac1 modification in addition to modulation of Rac1 activity by bt-15d-PGJ2 was observed in cultured ECs. In addition, decreased EC migration and cell spreading were observed in response to the electrophile. These results demonstrate for the first time that Rac1 is a target for 15d-PGJ2 in ECs, and suggest that Rac1 modification by electrophiles such as 15d-PGJ2 may alter redox signaling and EC function.
authors: Center for Free Radical Biology and Department of Pathology, University of Alabama at Birmingham, AL, USA.
authors: S B
authors: SB
authors: Wall
authors: Center for Free Radical Biology and Department of Pathology, University of Alabama at Birmingham, AL, USA.
authors: J-Y
authors: JY
authors: Oh
authors: Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, NC, USA.
authors: L
authors: L
authors: Mitchell
authors: Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, AL, USA.
authors: A H
authors: AH
authors: Laube
authors: Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, NC, USA.
authors: S L
authors: SL
authors: Campbell
authors: Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, AL, USA.
authors: M B
authors: MB
authors: Renfrow
authors: Center for Free Radical Biology and Department of Pathology, University of Alabama at Birmingham, AL, USA. Electronic address: landar@uab.edu.
authors: A
authors: A
authors: Landar
mesh_terms: D000818:Animals; D001011:Aorta / Q000166:cytology / Q000378:metabolism; D001710:Biotin / Q000737:chemistry; D002417:Cattle; D002465:Cell Movement; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D015870:Gene Expression; D010446:Peptide Fragments / Q000032:analysis; D061251:Primary Cell Culture; D015230:Prostaglandin D2 / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D059748:Proteolysis; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015398:Signal Transduction; D020830:rac1 GTP-Binding Protein / Q000737:chemistry / Q000378:metabolism*
chemical_list: C097240:15-deoxy-delta(12,14)-prostaglandin J2; D010446:Peptide Fragments; D011994:Recombinant Proteins; D001710:Biotin; D020830:rac1 GTP-Binding Protein; D015230:Prostaglandin D2
keywords: Cyclopentenone; Electrophile responsive proteome; Oxidative post-translational modification; Redox signaling; Rho GTPase; Thiol