title: 11-Deoxy,16,16-dimethyl prostaglandin E2 induces specific proteins in association with its ability to protect against oxidative stress.
abstract: Prostaglandins (PGs) act locally to maintain cellular homeostasis and stimulate stress response signaling pathways. These cellular effects are diverse and are tissue-dependent. PGE(2), and the synthetic analogue, 11-deoxy,16,16-dimethyl PGE(2) (DDM-PGE(2)), protect renal proximal tubular epithelial (LLC-PK1) cells against cellular injury induced by the potent nephrotoxic and nephrocarcinogenic metabolite of hydroquinone, 2,3,5-tris-(glutathion-S-yl)hydroquinone. Although this cytoprotective response (in LLC-PK1 cells) is mediated through a thromboxane or thromboxane-like receptor coupled to AP-1 signaling pathways, the mechanism of cytoprotection is unknown. In this study, we utilized HPLC-electrospray ionization tandem mass spectrometric (ESI MS/MS) and matrix-assisted laser desorption ionization time-of-flight mass spectrometric (MALDI TOF) analysis of proteins isolated from DDM-PGE(2)-stimulated LLC-PK1 cells to identify candidate cytoprotective proteins. DDM-PGE(2) selectively stimulated the synthesis of several proteins in LLC-PK1 cells. Peptide sequencing by ESI-MS/MS of in-gel tryptic protein digests revealed the identity of eight proteins: endothelial actin binding protein, myosin, elongation factor 2 (EF-2), elongation factor 1alpha-1 (EF-1alpha), heat shock protein 90beta (HSP90beta), glucose-regulated protein 78 (GRP 78), membrane-organizing extension spike protein, and actin. Both ESI-MS/MS and MALDI-MS analysis resulted in the same protein identification. Western analysis confirmed the temporal induction of the majority of these proteins, including EF-2, EF-1alpha, HSP90beta, GRP78, and actin. The collective expression of these proteins suggests that DDM-PGE(2)-mediated cytoprotection may involve alterations in cytoskeletal organization and/or stimulation of an endoplasmic reticulum (ER) stress response. The present studies provide insights into potential downstream targets of PG signaling.
authors.affiliation: Center for Molecular and Cellular Toxicology, Division of Pharmacology and Toxicology, College of Pharmacy, The University of Texas at Austin, Austin, Texas 78712, USA.
authors.forename: Kelly M
authors.initials: KM
authors.lastname: Towndrow
authors.forename: Zhe
authors.initials: Z
authors.lastname: Jia
authors.forename: Herng-Hsiang
authors.initials: HH
authors.lastname: Lo
authors.forename: Maria D
authors.initials: MD
authors.lastname: Person
authors.forename: Terrence J
authors.initials: TJ
authors.lastname: Monks
authors.forename: Serrine S
authors.initials: SS
authors.lastname: Lau
mesh_terms: D015064:16,16-Dimethylprostaglandin E2 / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology*; D000595:Amino Acid Sequence; D000818:Animals; D019610:Cytoprotection / Q000187:drug effects / Q000502:physiology; D030562:Databases, Protein; D000091342:Endoplasmic Reticulum Chaperone BiP; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D018374:LLC-PK1 Cells; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D020648:Peptide Elongation Factor 1 / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry; D014176:Protein Biosynthesis; D011506:Proteins / Q000302:isolation & purification* / Q000378:metabolism; D020539:Sequence Analysis, Protein; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D013552:Swine; D014357:Trypsin / Q000378:metabolism
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D020648:Peptide Elongation Factor 1; D010446:Peptide Fragments; D011506:Proteins; D014357:Trypsin; D015064:16,16-Dimethylprostaglandin E2