The Post-Transcriptional Regulation of Antioxidant Enzymes

A study was conducted exploring the post-transcriptional regulation of four antioxidant enzymes: glutathione peroxidase 1 (GPx1), manganese superoxide dismutase (MnSOD), glutathione peroxidase 4 (GPx4), and thioredoxin reductase 1 (TrxR1). Glutathione peroxidase activity was analyzed in clinical samples from imatinib-treated patients with chronic myelogenous leukemia (CML), which is characterized by the presence of Bcr-Abl. Studies were also conducted in the following human cell lines: KU812 and MEG-01 (CML), LNCaP (prostate cancer), and GM10832 (immortal B lymphocyte). Breakpoint Cluster Region-Abelson (Bcr- Abl) and Mammalian Target of Rapamycin (mTOR) were pharmacologically inhibited because their expression was expected to inhibit the protein levels of the selected antioxidants. Levels of antioxidant protein and activity were measured in cell lines treated with imatinib or rapamycin, the latter of which inhibits mTOR.;Glutathione peroxidase activity was found to increase in 4 of 7 CML patient sample sets, which was recapitulated in the two CML cell lines utilized for this study, in which MnSOD and TrxR1 were also increased in response to imatinib treatment. The increase of these three antioxidant proteins occurred only in CML cell lines. The ectopic expression of Bcr-Abl in the prostate cancer cell line LNCaP significantly decreased GPx1 protein; however, MnSOD and TrxR1 were unaltered. This indicates that GPx1 is potentially inhibited by Bcr-Abl, while MnSOD and TrxR1 could be increased in response to the intracellular effects of imatinib. In imatinib-treated CML cells, GPx1 and MnSOD activity levels increased only marginally in comparison to protein levels. This indicates that GPx1 and MnSOD may be post-translationally modified and that their activity is regulated independently of the post-transcriptional regulation that increased protein levels.;Treatment of both CML and non-CML cell lines with rapamycin, which inhibits mTOR, significantly increased protein levels for GPx1, GPx4, and TrxR1, suggesting that the posttranscriptional regulation of these proteins is dependent on the control of translation by mTOR and its downstream targets. These are all members of the selenoprotein family, the translation of which is dependent on the recoding of the selenocysteine codon by a selenocysteine insertion sequence (SECIS) found in the 3’-UTR of selenoprotein mRNA. The increase in these antioxidant proteins by mTOR inhibition illustrates a potential route by which selenoprotein translation may be regulated independent of selenium availability. Analysis of the effect of mTOR inhibition on a reporter construct dependent on the GPx1 SECIS element for expression did not reveal a difference in activity in the presence of rapamycin. This suggests that the mTOR-dependent translation of GPx1, GPx4, and TrxR1 may be via translation initiation and/or polypeptide chain elongation.