Subcellular Localization of Thioredoxin/Thioredoxin Reductase System—A Missing Link in Endoplasmic Reticulum Redox Balance
Abstract
:1. Introduction
2. Results
2.1. In Silico Analysis of Trx/TrxR Localization with Different Prediction Tools
2.2. Activity of TrxR in Subcellular Compartments of Rat Liver
2.3. Expression of Trx/TrxR Isoforms in Subcellular Compartments of Rat Liver
2.4. None of the Trx/TrxR Isoforms Co-Localizes with ER Marker Protein Grp94
2.5. ER-Targeted Expression of Trx1/TrxR1 in HeLa Cells Severely Compromised Cell Viability and Induced Apoptotic Cell Death
3. Discussion
4. Materials and Methods
4.1. In Silico Analysis
4.2. Animals
4.3. Preparation of Subcellular Fractions from Rat Liver Tissue
4.4. Measurement of Thioredoxin Reductase Activity
4.5. Sodium Dodecyl Sulphate Polyacrilamide Gel Electrophoresis (SDS-PAGE) and Western Blot Analysis
4.6. Cell Culture and Maintenance
4.7. Immunofluorescent Analysis of Human Fibroblast and HeLa Cells
4.8. Brightfield Microscopy of HeLa Cells
4.9. Plasmid Design and Construction
4.10. Transient Co-Transfection of HeLa Cells
4.11. Cell Viability Assay
4.12. Statistical Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Isoform | PSORT II | Predotar | Cello | MultiLoc2 | yLOC-HighRes | LocTree3 | DeepLoc-1.0 |
---|---|---|---|---|---|---|---|
Trx1 isoform_1 | - | 1% | 0.091 | 0% | 0.0% | - | 0.02% |
Trx1 isoform_2 | - | 1% | 0.091 | 0% | 0.0% | - | 9.20% |
Trx2_isoform 1 | - | 1% | 0.015 | 0% | 0.0% | - | 0.00% |
Trx2_isoform 2 † | 4.3% | - | 0.031 | 0% | 0.0% | - | 0.19% |
Trx2_isoform 3 † | 4.3% | 1% | 0.059 | 0% | 0.0% | - | 0.06% |
TrxR1_isoform 1 | - | 1% | 0.031 | 0% | 0.0% | - | 1.70% |
TrxR1_isoform 2 | - | 1% | 0.031 | 0% | 0.0% | - | 0.00% |
TrxR1_isoform 3 | - | 1% | 0.020 | 0% | 0.0% | - | 0.26% |
TrxR1_isoform 4 | - | 1% | 0.034 | 0% | 0.0% | - | 0.37% |
TrxR1_isoform 5 | - | 1% | 0.030 | 0% | 0.0% | - | 1.25% |
TrxR1_isoform 6 | - | 6% | 0.030 | 1% | 1.6% | - | 0.04% |
TrxR1_isoform 7 | - | 1% | 0.033 | 0% | 0.0% | - | 1.63% |
TrxR1_isoform 8 † | - | 1% | 0.022 | 1% | 0.0% | - | 0.00% |
TrxR1_isoform 9 † | - | 1% | 0.033 | 0% | 0.0% | - | 1.02% |
TrxR1_isoform 10 † | - | 1% | 0.034 | 0% | 0.2% | - | 1.78% |
TrxR1_isoform 11 † | - | 1% | 0.030 | 0% | 0.0% | - | 1.69% |
TrxR1_isoform 12 † | - | 1% | 0.038 | 0% | 0.0% | - | 0.70% |
TrxR1_isoform 13 † | - | 1% | 0.032 | 0% | 0.0% | - | 1.62% |
TrxR2_isoform 1 | - | 1% | 0.026 | 0% | 0.0% | - | 0.00% |
TrxR2_isoform 2 | - | 1% | 0.026 | 0% | 0.0% | - | 0.56% |
TrxR2_isoform 3 | - | 1% | 0.040 | 0% | 0.0% | - | 1.25% |
TrxR2_isoform 4 | - | 1% | 0.030 | 0% | 0.0% | - | 0.49% |
TrxR2_isoform 5 † | - | 1% | 0.026 | 0% | 0.0% | - | 2.11% |
TrxR2_isoform 6 † | - | 1% | 0.026 | 0% | 0.0% | - | 1.15% |
TrxR2_isoform 7 † | - | 1% | 0.025 | 0% | 0.0% | - | 0.00% |
TrxR2_isoform 8 † | - | 1% | 0.022 | 0% | 0.0% | - | 0.00% |
TrxR3_isoform 1 | 4.3% | 1% | 0.037 | 0% | 0.0% | - | 0.98% |
TrxR3_isoform 2 † | 4.3% | - | 0.037 | 0% | 0.0% | - | 0.90% |
TrxR3_isoform 3 † | 4.3% | 1% | 0.029 | 0% | 0.0% | - | 0.60% |
Localization | PSORT II | Predotar | Cello | MultiLoc2 | yLOC-HighRes | LocTree3 | DeepLoc-1.0 |
---|---|---|---|---|---|---|---|
Plasma membrane | - | - | 0.122 | 0% | 0.0% | - | 1.79% |
Endoplasmic reticulum | 4.3% | 1% | 0.029 | 0% | 0.0% | - | 0.60% |
Extracellular | - | - | 0.253 | 0% | 0.0% | - | 1.70% |
Lysosome | - | - | 0.026 | 0% | 0% | - | 0.93% |
Golgi | - | - | 0.021 | 0% | 0.0% | - | 0.35% |
Peroxysome | 4.3% | - | 0.271 | 30% | 2.6% | - | 16.43% |
Mitochondria | 17.4% | 1% | 1.401 | 10% | 0.8% | - | 11.35% |
Cytosol | 43.5% | - | 1.248 | 58% | 96.6% | 100% | 57.91% |
Nucleus | 21.7% | - | 0.735 | 2% | 0.0% | - | 4.25% |
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Veszelyi, K.; Czegle, I.; Varga, V.; Németh, C.E.; Besztercei, B.; Margittai, É. Subcellular Localization of Thioredoxin/Thioredoxin Reductase System—A Missing Link in Endoplasmic Reticulum Redox Balance. Int. J. Mol. Sci. 2024, 25, 6647. https://doi.org/10.3390/ijms25126647
Veszelyi K, Czegle I, Varga V, Németh CE, Besztercei B, Margittai É. Subcellular Localization of Thioredoxin/Thioredoxin Reductase System—A Missing Link in Endoplasmic Reticulum Redox Balance. International Journal of Molecular Sciences. 2024; 25(12):6647. https://doi.org/10.3390/ijms25126647
Chicago/Turabian StyleVeszelyi, Krisztina, Ibolya Czegle, Viola Varga, Csilla Emese Németh, Balázs Besztercei, and Éva Margittai. 2024. "Subcellular Localization of Thioredoxin/Thioredoxin Reductase System—A Missing Link in Endoplasmic Reticulum Redox Balance" International Journal of Molecular Sciences 25, no. 12: 6647. https://doi.org/10.3390/ijms25126647