Document Detail

The expression of Exonuclease III from E. coli in mitochondria of breast cancer cells diminishes mitochondrial DNA repair capacity and cell survival after oxidative stress.
MedLine Citation:
PMID:  12713808     Owner:  NLM     Status:  MEDLINE    
The ability to sensitize cancer cells to radiation would be highly beneficial for successful cancer treatment. One mode of action for ionizing radiation is the induction of cell death through infliction of extensive oxidative damage to cellular DNA, including mitochondrial DNA (mtDNA). The ability of cells to repair mtDNA and otherwise maintain the integrity of their mitochondria is vital for protection of the cells against oxidative damage. Because efficient repair of oxidative damage in mtDNA may play a crucial role in cancer cell resistance, interference with this repair process could be an effective way to achieve a radiation sensitive phenotype in otherwise resistant cancer cells. Successful repair of DNA is achieved through a precise and highly regulated multistep process. Expression of excessive amounts of one of the repair enzymes may cause an imbalance of the whole repair system and lead to the loss of repair efficiency. To study the effects of changing mtDNA repair capacity on overall cell survival following oxidative stress, we expressed a bacterial repair enzyme, Exonuclease III (ExoIII) containing the mitochondrial targeting signal of manganese superoxide dismutase, in a human malignant breast epithelial cell line, MDA-MB-231. Following transfection, specific exonuclease activity was found in mitochondrial extracts. In order to examine the effects on repair of oxidative damage in mtDNA, cells were exposed to the enzyme xanthine oxidase and its substrate hypoxanthine. mtDNA repair was evaluated using quantitative Southern blot analysis. The results revealed that cells expressing ExoIII in mitochondria are deficient in mtDNA repair when compared with control cells that express ExoIII without MTS. This diminished mtDNA repair capacity rendered MDA-MB-231 cells more sensitive to oxidative damage, which resulted in a decrease in their long-term survival following oxidative stress.
Inna N Shokolenko; Mikhail F Alexeyev; Fredika M Robertson; Susan P LeDoux; Glenn L Wilson
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  DNA repair     Volume:  2     ISSN:  1568-7864     ISO Abbreviation:  DNA Repair (Amst.)     Publication Date:  2003 May 
Date Detail:
Created Date:  2003-04-25     Completed Date:  2004-01-14     Revised Date:  2007-11-15    
Medline Journal Info:
Nlm Unique ID:  101139138     Medline TA:  DNA Repair (Amst)     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  471-82     Citation Subset:  IM    
Department of Cell Biology, University of South Alabama College of Medicine, Mobile, AL 36688, USA.
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MeSH Terms
Blotting, Southern
Blotting, Western
Breast Neoplasms / metabolism*
Cell Line, Tumor
Cell Survival
DNA Repair*
DNA, Mitochondrial / metabolism*
Dose-Response Relationship, Drug
Escherichia coli / enzymology*
Exodeoxyribonucleases / biosynthesis*
Mitochondria / metabolism*
Oxidative Stress*
Polymerase Chain Reaction
Radiation, Ionizing
Reactive Oxygen Species
Subcellular Fractions / metabolism
Time Factors
Xanthine Oxidase / metabolism
Grant Support
Reg. No./Substance:
0/DNA, Mitochondrial; 0/Reactive Oxygen Species; EC Oxidase; EC 3.1.-/Exodeoxyribonucleases; EC III

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