Transfection of mGSTA4 in HL-60 cells protects against 4-hydroxynonenal-induced apoptosis by inhibiting JNK-mediated signaling.
The mammalian alpha-class glutathione S-transferase (GST) isozymes mGSTA4-4, rGSTA4-4, and hGSTA4-4 are known to utilize 4-hydroxynonenal (4HNE) as a preferred substrate. During the present studies, we have examined the effect of transfecting human myeloid HL-60 cells with mGSTA4, on 4-HNE-induced apoptosis and the associated signaling mechanisms. Results of these studies show that treatment of the wild-type or vector-only-transfected HL-60 cells with 20 microM 4-HNE caused apoptosis within 2 h. The cells transfected with mGSTA4 did not undergo apoptosis under these conditions even after 4 h. In the wild-type and vector-transfected cells, apoptosis was preceded by JNK activation and c-Jun phosphorylation within 30 min, and an increase in AP-1 binding within 2 h of treatment with 20 microM 4-HNE. In mGSTA4-transfected cells, JNK activation and c-Jun phosphorylation were observed after 1 h, and increased AP-1 binding was observed after 8 h under these conditions. In the control cells, 20 microM 4-HNE caused caspase 3 activation and poly(ADP-ribose) polymerase cleavage within 2 h, while in mGSTA4-transfected cells, a lesser degree of these effects was observed even after 8 h. Transfection with mGSTA4 also provided protection to the cells from 4-HNE and doxorubicin cytotoxicity (1.6- and 2.6-fold, respectively). These results show that 4-HNE mediates apoptosis through its effects on JNK and caspase 3, and that 4-HNE metabolizing GST isozyme(s) may be important in the regulation of this pathway of oxidative-stress-induced apoptosis.
Cheng
J Z
JZ
Department of Human Biological Chemistry and Genetics, UTMB, Galveston, Texas 77555-1067, USA.
Singhal
S S
SS
Sharma
A
A
Saini
M
M
Yang
Y
Y
Awasthi
S
S
Zimniak
P
P
Awasthi
Y C
YC
D000447:Aldehydes / Q000494:pharmacology*; D017209:Apoptosis; D015152:Blotting, Northern; D015153:Blotting, Western; D002469:Cell Separation; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D005982:Glutathione Transferase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D018922:HL-60 Cells; D006801:Humans; D020287:In Situ Nick-End Labeling; D007527:Isoenzymes; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D018384:Oxidative Stress; D015398:Signal Transduction; D013997:Time Factors; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014162:Transfection
D000447:Aldehydes; D015853:Cysteine Proteinase Inhibitors; D007527:Isoenzymes; D018808:Transcription Factor AP-1; D005982:Glutathione Transferase; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; C027576:4-hydroxy-2-nonenal