Thioredoxin as a molecular target of cyclopentenone prostaglandins.

Abstract

Prostaglandin (PG) D2, a major cyclooxygenase product in a variety of tissues and cells, readily undergoes dehydration to yield the bioactive cyclopentenone-type PGs of the J2 series, such as 15-deoxy-Delta12,14-PGJ2 (15d-PGJ2). We have shown previously that 15d-PGJ2 is a potent electrophile that causes intracellular oxidative stress and redox alteration in human neuroblastoma SH-SY5Y cells. In the present study, based on the observation that the electrophilic center of 15d-PGJ2 was involved in the pro-oxidant effect, we investigated the role of thioredoxin 1 (Trx), an endogenous redox regulator, against 15d-PGJ2-induced oxidative cell injury. It was observed that the 15d-PGJ2-induced oxidative stress was significantly suppressed by the Trx overexpression. In addition, the treatment of SH-SY5Y cells with biotinylated 15d-PGJ2 resulted in the formation of a 15d-PGJ2-Trx adduct, indicating that 15d-PGJ2 directly modified the endogenous Trx in the cells. To further examine the mechanism of the 15d-PGJ2 modification of Trx, human recombinant Trx treated with 15d-PGJ2 was analyzed by mass spectrometry. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry analysis of the 15d-PGJ2-treated human recombinant Trx demonstrated the addition of one molecule of 15d-PGJ2 per protein molecule. Moreover, the electrospray ionization-liquid chromatography/mass spectrometry/mass spectrometry analysis identified two cysteine residues, Cys-35 and Cys-69, as the targets of 15d-PGJ2. These residues may represent the direct sensors of the electrophilic PGs that induce the intracellular redox alteration and neuronal cell death.

• Authors: Takahiro Shibata, Takaaki Yamada, Takeshi Ishii, Shigenori Kumazawa, Hajime Nakamura, Hiroshi Masutani, Junji Yodoi, Koji Uchida
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 278 Issue 28 Pg. 26046-54 (Jul 11 2003) ISSN: 0021-9258 [Print] United States
• PMID: 12709421 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• 15-deoxy-delta(12,14)-prostaglandin J2
• Cyclopentanes
• Oxidants
• Peptides
• Prostaglandins
• Recombinant Proteins
• Thioredoxins
• cyclopentenone
• Prostaglandin D2
• Oxygen

Topics

• Amino Acid Sequence
• Biotinylation
• Cell Death
• Cyclopentanes (chemistry)
• Dose-Response Relationship, Drug
• Electrophoresis, Polyacrylamide Gel
• Enzyme-Linked Immunosorbent Assay
• Flow Cytometry
• Humans
• Immunoblotting
• Models, Chemical
• Molecular Sequence Data
• Neurons (pathology)
• Oxidants (pharmacology)
• Oxidation-Reduction
• Oxidative Stress
• Oxygen (metabolism)
• Peptides (chemistry)
• Prostaglandin D2 (analogs & derivatives, metabolism)
• Prostaglandins (chemistry, metabolism)
• Protein Binding
• Recombinant Proteins (chemistry, metabolism)
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
• Thioredoxins (metabolism)
• Transfection
• Tumor Cells, Cultured