Abstract
Glutathione transferase P1-1 (GSTP1-1) plays crucial roles in cancer chemoprevention and chemoresistance and is a key target for anticancer drug development. Oxidative stress or inhibitor-induced GSTP1-1 oligomerization leads to the activation of stress cascades and apoptosis in various tumor cells. Therefore, bivalent glutathione transferase (GST) inhibitors with the potential to interact with GST dimers are been sought as pharmacological and/or therapeutic agents. Here we have characterized GSTP1-1 oligomerization in response to various endogenous and exogenous agents. Ethacrynic acid, a classic GSTP1-1 inhibitor, 4-hydroxy-nonenal, hydrogen peroxide, and diamide all induced reversible GSTP1-1 oligomerization in Jurkat leukemia cells through the formation of disulphide bonds involving Cys47 and/or Cys101, as suggested by reducing and nonreducing SDS-polyacrylamide gel electrophoresis analysis of cysteine to serine mutants. Remarkably, the electrophilic prostanoid 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) induced irreversible GSTP1-1 oligomerization, specifically involving Cys101, a residue present in the human but not in the murine enzyme. 15d-PGJ2-induced GSTP1-1 cross-linking required the prostaglandin (PG) dienone structure and was associated with sustained c-Jun NH2-terminal kinase activation and induction of apoptosis. It is noteworthy that 15d-PGJ2 elicited GSTP1-1 cross-linking in vitro, a process that could be mimicked by other dienone cyclopentenone PG, such as Δ12-PGJ2, and by the bifunctional thiol reagent dibromobimane, suggesting that cyclopentenone PG may be directly involved in oligomer formation. Remarkably, Δ12-PGJ2-induced oligomeric species were clearly observed by electron microscopy showing dimensions compatible with GSTP1-1 tetramers. These results provide the first direct visualization of GSTP1-1 oligomeric species. Moreover, they offer novel strategies for the modulation of GSTP1-1 cellular functions, which could be exploited to overcome its role in cancer chemoresistance.
Footnotes
This work was supported by the Spanish Ministry of Science and Innovation [Grants SAF-2006-03489, SAF-2009-11642, BFU2008-00666, BFU-2009-08977, SAF-2008-00451]; Instituto de Salud Carlos III [Grants RD07/0064/0007, RD06/0020/1001]; Human Frontiers Science Program [Grant RGP39/2008]; and the Autonomous Region of Madrid [Grant CAM S-BIO-0214-2006].
Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
doi:10.1124/mol.110.065391.
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ABBREVIATIONS:
- GST
- glutathione transferase
- GSTP1-1
- glutathione transferase isoform P1-1
- JNK
- c-Jun NH2-terminal kinase
- cyPG
- cyclopentenone prostaglandin(s)
- 15d-PGJ2
- 15-deoxy-Δ12,14-prostaglandin J2
- 15d-PGJ2-B
- 15d-PGJ2-biotin
- PGA1-B
- PGA1-biotin
- PAGE
- polyacrylamide gel electrophoresis
- HRP
- horseradish peroxidase
- ECL
- enhanced chemiluminescence
- RMC
- rat mesangial cell
- PG
- prostaglandin
- DTT
- dithiothreitol
- PCR
- polymerase chain reaction
- 2D
- two-dimensional
- HNE
- 4-hydroxy-2-nonenal.
- Received April 6, 2010.
- Accepted July 13, 2010.
- Copyright © 2010 The American Society for Pharmacology and Experimental Therapeutics
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