MnSOD and Cyclin B1 Coordinate a Mito-Checkpoint during Cell Cycle Response to Oxidative Stress
Abstract
:1. Introduction
2. Material and Methods
2.1. Cell Line and Culture Conditions
2.2. Cell Viability Assay
2.3. Flow Cytometry Assay of DNA Content
2.4. Sub-Cellular Fractionation
2.5. Immunoblotting Assay
2.6. MnSOD Activity Gel Assay
2.7. Complex I, II, IV and Aconitase Activity Assays
2.8. Immune-Staining and Confocal Microscopy
2.9. Immuno-Gold Staining and Electron Microscopy
2.10. Mitochondria-Targeted Expression of Human Cyclin B1
2.11. Dihydroethidium (DHE) for ROS Levels and Mitochondrial Membrane Potential Assays
3. Results
3.1. Cyclin B1 Translocates to Mitochondria Following Oxidative Stress Affecting Mitochondrial Function
3.2. Cyclin B1 Mitochondria and Nuclear Cross-Talk Was Associated with Oxidative Stress Induced Delay and Restart of the Cell Cycle
3.3. Cyclin B1 Mitochondrial Translocation Perturbs Mitochondria Functions
3.4. Mitochondria-Targeted Cyclin B1 Overexpression Mimics Many of the Cellular Properties Observed Following Oxidative Stress
4. Discussion
5. Conclusions
Acknowledgments
Author Contributions
Conflicts of Interest
Abbreviations
AKAP | A-kinase anchor proteins |
BCl-2 | B-cell lymphoma 2 |
CCCP | Carbonyl cyanide m-chlorophenyl hydrazone |
CDK1 | Cyclin-dependent kinase 1 |
CuZnSOD | Copper Zinc Superoxide Dismutase |
DHE | Dihydroethidium |
DMEM | Dulbecco’s modified Eagle’s medium |
EPR | Electron paramagnetic resonance |
ERK | Extracellular Signal-regulated Kinase |
FBS | Fetal Bovine Serum |
GAPDH | Glyceraldehyde-3-phospahte dehydrogenase |
Gy | Gray |
h | hour(s) |
HBSS | Hanks buffer salt solution |
IR | Ionizing radiation |
JNK | c-Jun NH2-terminal kinase |
MAP | Mitogen-activated protein |
MCF10A | Human Mammary Epithelial cells |
MFI | Mean fluorescence intensity |
MnSOD | Manganese Superoxide Dismutase |
NADH | Nicotinamide adenine dinucleotide |
NBT | Nitroblue tetrazolium |
NHF | Normal Human Skin Fibroblast |
PBS | Phosphate Buffered Saline |
PDT | Photodynamic therapy |
PI | Propidium Iodide |
PMT | photomultiplier |
ROS | Reactive oxygen species |
SDS-PAGE | Sodium dodecyl sulfate polyacrylamide gel electrophoresis |
SRC | SRC proto-oncogene, non-receptor tyrosine kinase |
TCA | Tricarboxylic acid cycle |
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Kalen, A.L.; Ahmad, I.M.; Abdalla, M.Y.; O’Malley, Y.Q.; Goswami, P.C.; Sarsour, E.H. MnSOD and Cyclin B1 Coordinate a Mito-Checkpoint during Cell Cycle Response to Oxidative Stress. Antioxidants 2017, 6, 92. https://doi.org/10.3390/antiox6040092
Kalen AL, Ahmad IM, Abdalla MY, O’Malley YQ, Goswami PC, Sarsour EH. MnSOD and Cyclin B1 Coordinate a Mito-Checkpoint during Cell Cycle Response to Oxidative Stress. Antioxidants. 2017; 6(4):92. https://doi.org/10.3390/antiox6040092
Chicago/Turabian StyleKalen, Amanda L., Iman M. Ahmad, Maher Y. Abdalla, Yunxia Q. O’Malley, Prabhat C. Goswami, and Ehab H. Sarsour. 2017. "MnSOD and Cyclin B1 Coordinate a Mito-Checkpoint during Cell Cycle Response to Oxidative Stress" Antioxidants 6, no. 4: 92. https://doi.org/10.3390/antiox6040092
APA StyleKalen, A. L., Ahmad, I. M., Abdalla, M. Y., O’Malley, Y. Q., Goswami, P. C., & Sarsour, E. H. (2017). MnSOD and Cyclin B1 Coordinate a Mito-Checkpoint during Cell Cycle Response to Oxidative Stress. Antioxidants, 6(4), 92. https://doi.org/10.3390/antiox6040092