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Modulation of glucose-6-phosphate dehydrogenase activity and expression is associated with aryl hydrocarbon
G C Yeh1, P J Daschner, J Lopaczynska
1Cellular Defense and Carcinogenesis Section, Basic Research Laboratory, NCI at Frederick, National Institutes of Health, Frederick, Maryland 21702, USA. yeh@ncifcrf.gov
Abstract:
The mutagenic effect of environmental carcinogens has been well documented in animal models and in human studies but the mechanisms involved in preventing carcinogen insult have not been fully elucidated. In this study we examined the molecular and biochemical changes associated with carcinogen resistance in a series of aryl hydrocarbon-resistant MCF-7 cell lines developed by exposure to benzo[a]pyrene (BP). The cell lines were designated as AH(R40), AH(R100), and AH(R200) to denote their increasing fold resistance to BP compared with wild type cells. These cell lines were also resistant to another aryl hydrocarbon (AH), dimethylbenz[a]anthracene, but not to pleiotropic drugs (doxorubicin, vinblastine, and taxol). The resistant cell lines showed an increase in the level of the primary intracellular antioxidant, reduced glutathione, corresponding to increasing AH resistance. However, there was no change in glutathione reductase activity. The generation of reduced glutathione requires NADPH, and we therefore examined the activity and expression of the rate-limiting enzyme in NADPH production, glucose-6-phosphate dehydrogenase (G6PD). An increase in G6PD specific activity was associated with increasing aryl hydrocarbon resistance. This was due to an increased expression of G6PD in resistant cells, which was demonstrated by increases in both protein and mRNA levels. However, there was no increase in the transcription rate of G6PD in the resistant cell lines, indicating that the increase G6PD expression is due to a post-transcriptional modulation, which was confirmed by actinomycin D chase experiments. These results demonstrate that modulation of G6PD expression and activity is an important mechanism in AH resistance.
Insights
Environmental carcinogen resistance in cells involves increased glucose-6-phosphate dehydrogenase (G6PD) expression. This post-transcriptional modulation enhances antioxidant defenses, crucial for cellular protection against harmful compounds.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Mechanisms of cellular protection against environmental carcinogens are not fully understood.
- Aryl hydrocarbon (AH) resistance in cancer cells is a complex phenomenon.
- Previous studies highlight the role of antioxidants in mitigating carcinogen damage.
Purpose of the Study:
- To investigate the molecular and biochemical basis of aryl hydrocarbon resistance in MCF-7 cells.
- To identify key enzymes and pathways involved in cellular defense against benzo[a]pyrene (BP).
- To elucidate the regulatory mechanisms controlling enzyme expression in resistant cells.
Main Methods:
- Development of benzo[a]pyrene-resistant MCF-7 cell lines (AH(R40), AH(R100), AH(R200)).
- Measurement of intracellular reduced glutathione levels and glutathione reductase activity.
- Assay of glucose-6-phosphate dehydrogenase (G6PD) activity, protein, and mRNA levels.
- Analysis of G6PD transcription rates and post-transcriptional regulation using actinomycin D.
Main Results:
- Resistant cell lines exhibited cross-resistance to another AH but not to unrelated drugs.
- Increased levels of reduced glutathione correlated with increasing AH resistance.
- Elevated G6PD specific activity, protein, and mRNA levels were observed in resistant cells.
- G6PD expression increased post-transcriptionally, not via enhanced transcription.
Conclusions:
- Modulation of glucose-6-phosphate dehydrogenase (G6PD) expression is a key mechanism conferring aryl hydrocarbon resistance.
- Post-transcriptional regulation of G6PD plays a significant role in cellular adaptation to carcinogen exposure.
- Increased G6PD activity supports enhanced antioxidant capacity, protecting cells from carcinogen insult.