title: Oligomeric state regulated trafficking of human platelet-activating factor acetylhydrolase type-II.
abstract: The intracellular enzyme platelet-activating factor acetylhydrolase type-II (PAFAH-II) hydrolyzes platelet-activating factor and oxidatively fragmented phospholipids. PAFAH-II in its resting state is mainly cytoplasmic, and it responds to oxidative stress by becoming increasingly bound to endoplasmic reticulum and Golgi membranes. Numerous studies have indicated that this enzyme is essential for protecting cells from oxidative stress induced apoptosis. However, the regulatory mechanism of the oxidative stress response by PAFAH-II has not been fully resolved. Here, changes to the oligomeric state of human PAFAH-II were investigated as a potential regulatory mechanism toward enzyme trafficking. Native PAGE analysis in vitro and photon counting histogram within live cells showed that PAFAH-II is both monomeric and dimeric. A Gly-2-Ala site-directed mutation of PAFAH-II demonstrated that the N-terminal myristoyl group is required for homodimerization. Additionally, the distribution of oligomeric PAFAH-II is distinct within the cell; homodimers of PAFAH-II were localized to the cytoplasm while monomers were associated to the membranes of the endoplasmic reticulum and Golgi. We propose that the oligomeric state of PAFAH-II drives functional protein trafficking. PAFAH-II localization to the membrane is critical for substrate acquisition and effective oxidative stress protection. It is hypothesized that the balance between monomer and dimer serves as a regulatory mechanism of a PAFAH-II oxidative stress response.
authors.affiliation: Department of Chemistry & Biochemistry, University of Delaware, Newark, DE 19716, USA.
authors.forename: Elizabeth S
authors.initials: ES
authors.lastname: Monillas
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, DE 19716, USA.
authors.forename: Jeffrey L
authors.initials: JL
authors.lastname: Caplan
authors.affiliation: Department of Chemistry & Biochemistry, University of Delaware, Newark, DE 19716, USA.
authors.forename: Anastasia F
authors.initials: AF
authors.lastname: Thévenin
authors.affiliation: Department of Chemistry & Biochemistry, University of Delaware, Newark, DE 19716, USA. Electronic address: bahnson@udel.edu.
authors.forename: Brian J
authors.initials: BJ
authors.lastname: Bahnson
mesh_terms: D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase / Q000737:chemistry / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008958:Models, Molecular; D018384:Oxidative Stress; D055503:Protein Multimerization / Q000502:physiology*; D021381:Protein Transport; D014018:Tissue Distribution
chemical_list: D043203:1-Alkyl-2-acetylglycerophosphocholine Esterase
keywords: Myristoylation; Oligomeric state; Oxidative stress; Peripheral membrane protein; Phospholipase A2; Trafficking