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Peplomycin-induced DNA repair synthesis in permeable mouse ascites sarcoma cells
B Zhang1, S Watanabe, K Akiyama
1Department of Biochemistry, Okayama University Medical School, Japan.
Abstract:
DNA repair synthesis induced in permeable mouse ascites sarcoma cells by peplomycin, an antitumor antibiotic, was studied. Mouse ascites sarcoma (SR-C3H/He) cells were permeabilized with a low concentration of Triton X-100 in an isotonic condition. Permeable cells were treated with an appropriate concentration of peplomycin to introduce single-strand breaks in permeable cell DNA. DNA repair synthesis in peplomycin-treated permeable cells was measured by incubating the cells with four deoxynucleoside triphosphates in an appropriate buffer system. The DNA repair synthesis was enhanced by ATP and NaCl at near physiological concentrations. More than 90% of DNA synthesis in the present system depended on the peplomycin-treatment. The repair nature of the DNA synthesis was confirmed by a BrdUMP density shift technique. The repair patches were largely completed and ligated in the presence of ATP. Analyses using selective inhibitors for DNA polymerases showed that both DNA polymerase Beta and aphidicolin-sensitive DNA polymerases (DNA polymerase alpha and/or delta) were involved in the repair DNA synthesis.
Insights
This study shows peplomycin induces DNA repair synthesis in mouse sarcoma cells. ATP and NaCl enhance this repair, involving DNA polymerases Beta, alpha, and delta.
Area of Science:
- Molecular Biology
- Cancer Research
- Biochemistry
Background:
- DNA damage is a critical factor in cancer development and treatment.
- Understanding DNA repair mechanisms is essential for developing effective cancer therapies.
- Peplomycin is an antitumor antibiotic known to induce DNA strand breaks.
Purpose of the Study:
- To investigate DNA repair synthesis in mouse ascites sarcoma cells induced by peplomycin.
- To identify the conditions and enzymes involved in peplomycin-induced DNA repair.
Main Methods:
- Mouse ascites sarcoma cells were permeabilized using Triton X-100.
- Permeable cells were treated with peplomycin to induce DNA single-strand breaks.
- DNA repair synthesis was measured using deoxynucleoside triphosphates and confirmed via BrdUMP density shift technique.
Main Results:
- Peplomycin treatment induced significant DNA repair synthesis, with over 90% dependent on the treatment.
- Adenosine triphosphate (ATP) and sodium chloride (NaCl) at physiological concentrations enhanced DNA repair synthesis.
- DNA polymerase Beta and aphidicolin-sensitive DNA polymerases (alpha and/or delta) were implicated in the repair process.
Conclusions:
- Peplomycin effectively induces DNA repair synthesis in permeable mouse sarcoma cells.
- ATP and NaCl play crucial roles in facilitating DNA repair patch completion and ligation.
- The study highlights the involvement of multiple DNA polymerases in repairing peplomycin-induced DNA damage.