• Radiation Oncology Journal
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• 제11권2호
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• pp.219-225
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• 1993

각 방사선량의 조사에 따른 사람 정상임파구, 마우스 정상임파구, 그리고 EL-4마우스 백혈병 세포의 DNA double strand breaks 발생율에 대하여 비교분석 하였던 바 다음과 같은 결과를 얻었다. LPS와 PHA를 첨가하여 각각 배양한 마우스 정상임파구의 DNA double strand breaks의발생율을 strand scission factor의 기울기로 비교평가해 본 바 LPS를 첨가배양한 군이 DNA DSB가 더 많이 형성되었다(P<0.005). 이것으로 볼 때 DNA double strand breaks발생에 있어서는 B cells이 T cells보다 더 민감하였다. 또한 EL-4 백혈병 세포는 정상 임파구보다 유의하게 DNA DSB가 더 많이 형성된 것이 관찰되었다(p<0.005). 한편 사람 정상임파구와 마우스 정상임파구 사이의 intrinsic radiosensitivity는 시험관내 실험에서 비교적 유사한 kinetics를 나타냈으나, 마우스 정상임파구가 사람 정상임파구보다 DNA DSB수율이 더 낮았다. 그래서 방사선을 조사하고 3.5시간이 지난후에 $10\%$ DNA DSB발생에 필요한 선량을 두 군간에 비교평가해 본 바 마우스 정상임파구의 선량이 더 높게 나타났다. 이상의 실험결과를 정확하게 설명하기는 어 렵지만 아마도 환경적인 요인과 유전적인 요인 때문인 것으로 사료되었다. 그리고 본 연구결과는 방사선조사에 따른 임상반응의 이해와 cytometric assessment의 방사선 생물학적 parameter로 이용될 수 있을 것으로 판단되었다.

• Journal of Microbiology and Biotechnology
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• 제30권3호
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• pp.469-475
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• 2020

During meiosis I, programmed DNA double-strand breaks (DSBs) occur to promote chromosome pairing and recombination between homologs. In Saccharomyces cerevisiae, Mec1 and Tel1, the orthologs of human ATR and ATM, respectively, regulate events upstream of the cell cycle checkpoint to initiate DNA repair. Tel1^ATM and Mec1^ATR are required for phosphorylating various meiotic proteins during recombination. This study aimed to investigate the role of Tel1^ATM and Mec1^ATR in meiotic prophase via physical analysis of recombination. Tel1^ATM cooperated with Mec1^ATR to mediate DSB-to-single end invasion transition, but negatively regulated DSB formation. Furthermore, Mec1^ATR was required for the formation of interhomolog joint molecules from early prophase, thus establishing a recombination partner choice. Moreover, Mec1^ATR specifically promoted crossover-fated DSB repair. Together, these results suggest that Tel1^ATM and Mec1^ATR function redundantly or independently in all post-DSB stages.

• Journal of Microbiology and Biotechnology
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• 제25권7호
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• pp.1026-1035
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• 2015

Condensin is not only responsible for chromosome condensation, but is also involved in double-strand break (DSB) processing in the cell cycle. During meiosis, the condensin complex serves as a component of the meiotic chromosome axis, and mediates both proper assembly of the synaptonemal complex and DSB repair, in order to ensure proper homologous chromosome segregation. Here, we used the budding yeast Saccharomyces cerevisiae to show that condensin participates in a variety of chromosome organization processes and exhibits crucial molecular functions that contribute to meiotic recombination during meiotic prophase I. We demonstrate that Ycs4 is required for efficient DSB formation and establishing homolog bias at the early stage of meiotic prophase I, which allows efficient formation of interhomolog recombination products. In the Ycs4 meiosis-specific allele (ycs4S), interhomolog products were formed at substantial levels, but with the same reduction in crossovers and noncrossovers. We further show that, in prophase chromosomal events, ycs4S relieved the defects in the progression of recombination interactions induced as a result of the absence of Rec8. These results suggest that condensin is a crucial coordinator of the recombination process and chromosome organization during meiosis.

• Radiation Oncology Journal
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• 제13권2호
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• pp.113-120
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• 1995

Purpose : To investigate the effect of alkaloid fraction from Korean ginseng on radiation-induced DNA double strand breaks (dsb) formation and repair in murine lymphocytes Materials and Methods : We used the neutral filter elution technique to assay $^{60}Co\;{\gamma}$ ray-induced DNA double strand breaks formation and repair in C57BL/6 mouse spleen lymphocytes for evaluating the dose-response relationship in the presence of alkaloid fraction as a radioprotective agent. The lymphocytes were stimulated with Phytohemagglutinin (PHA, 2 u g/ml) to label $^3[H]-thymidine.$ Isotope-labelled lymphocytes in suspension were exposed to 100 Gy at $0^{\cdot}C$ in the alkaloid fraction-treated group and elution procedure was performed at PH 9.6. The extents of formation of radiation-induced DNA double strand breaks and repair were compared respectively via strand scission factor (SSF) and relative strand scission factor (RSSF). Results: Alkaloid fraction reduced the formation of double strand breaks with dose modification factor of 2 15, compared to control group Rejoining of DNA dsb appeared to take place via two components. The first fast component was completed within 20.4 minutes, but the second slow component was not completed until 220.2 minutes after irradiation. About $30\%$ of dsb formed by irradiation was ultimately unrejoined despite the administration of alkaloid fraction. The administration of alkaloid fraction had a great effect on the second slow component of repair; the half-time of fast component repair was not changed, but that of slow component was 621.8 minutes. Conclusion: Neutral filter elution assay Proved to be a very effective method to quantitate the extents of DNA dsb formation and its repair. By using this technique, we were able to evaluate the efficiency of alkaloid fraction from Korean ginseng as a valuable radioprotector. Alkaloid fraction can be used prophylactically to prevent or ameliorate the severe radiation damages in workers and neighbors around the atomic power plants. For more refined study, however, more advanced purification of alkaloid fraction wil be needed in the near future.

• Journal of Microbiology and Biotechnology
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• 제25권5호
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• pp.598-605
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• 2015

The cohesin complex holds sister chromatids together and prevents premature chromosome segregation until the onset of anaphase. Mcd1 (also known as Scc1), the α-kleisin subunit of cohesin, is a key regulatory subunit of the mitotic cohesin complex and is required for maintaining sister chromatid cohesion, chromosome organization, and DNA repair. We investigated the function of Mcd1 in meiosis by ectopically expressing Mcd1 during early meiotic prophase I in Saccharomyces cerevisiae. Mcd1 partially regulated the progression of meiotic recombination, sister chromatid separation, and nuclear division. DNA physical analysis during meiotic recombination showed that Mcd1 induced double-strand breaks (DSBs) but negatively regulated homologous recombination during DSB repair; Mcd1 expression delayed post-DSB stages, leading to inefficiencies in the DSB-to-joint molecule (JM) transition and subsequent crossover formation. These findings indicate that meiotic cells undergo Mcd1-mediated DSB formation during prophase I, and that residual Mcd1 could regulate the progression of JM formation during meiotic recombination.

• Journal of electromagnetic engineering and science
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• 제15권3호
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• pp.134-141
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• 2015

The effect of a 60 Hz time-varying electric field was studied using a facing-electrode device (FED) and a coplanar-electrode device (CED) for further investigation of the genotoxicity of 60 Hz time-varying magnetic field (MF) from preceding research. Neither a single 30-minute exposure to the CED or to the FED had any obvious biological effects such as DNA double strand break (DSB) and apoptosis in cancerous SCC25, and HeLa cells, normal primary fibroblast IMR90 cells, while exposures of 60 Hz time-varying MF led to DNA damage with induced electric fields much smaller than those used in this experiment. Nor did repetitive exposures of three days or a continuous exposure of up to 144 hours with the CED induce any DNA damage or apoptosis in either HeLa or IMR90 cells. These results imply that the solitary electric field produced by time-varying MF is not a major cause of DSBs or apoptosis in cancer or normal cells.

• Molecules and Cells
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• 제42권7호
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• pp.546-556
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• 2019

Protein phosphatase 4 (PP4) is a crucial protein complex that plays an important role in DNA damage response (DDR), including DNA repair, cell cycle arrest and apoptosis. Despite the significance of PP4, the mechanism by which PP4 is regulated remains to be elucidated. Here, we identified a novel PP4 inhibitor, protein phosphatase 4 inhibitory protein (PP4IP) and elucidated its cellular functions. PP4IP-knockout cells were generated using the CRISPR/Cas9 system, and the phosphorylation status of PP4 substrates (H2AX, KAP1, and RPA2) was analyzed. Then we investigated that how PP4IP affects the cellular functions of PP4 by immunoprecipitation, immunofluorescence, and DNA double-strand break (DSB) repair assays. PP4IP interacts with PP4 complex, which is affected by DNA damage and cell cycle progression and decreases the dephosphorylational activity of PP4. Both overexpression and depletion of PP4IP impairs DSB repairs and sensitizes cells to genotoxic stress, suggesting timely inhibition of PP4 to be indispensable for cells in responding to DNA damage. Our results identify a novel inhibitor of PP4 that inhibits PP4-mediated cellular functions and establish the physiological importance of this regulation. In addition, PP4IP might be developed as potential therapeutic reagents for targeting tumors particularly with high level of PP4C expression.

• BMB Reports
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• 제55권11호
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• pp.541-546
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• 2022

The repair of DNA double-strand breaks (DSBs) by homologous recombination (HR) is crucial for maintaining genomic integrity and is involved in numerous fundamental biological processes. Post-translational modifications by proteins play an important role in regulating DNA repair. Here, we report that the methyltransferase SET7 regulates HR-mediated DSB repair by methylating TIP60, a histone acetyltransferase and tumor suppressor involved in gene expression and protein stability. We show that SET7 targets TIP60 for methylation at K137, which facilitates DSB repair by promoting HR and determines cell viability against DNA damage. Interestingly, TIP60 demethylation is catalyzed by LSD1, which affects HR efficiency. Taken together, our findings reveal the importance of TIP60 methylation status by SET7 and LSD1 in the DSB repair pathway.

• 대한수의학회지
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• 제31권3호
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• pp.329-335
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• 1991

The filter elution technique was used to assay $^{60}Co$ $\gamma$ ray-induced DNA strand breaks(SB) in EL4 mouse leukemia cell and mouse spleen lymphocyte. The lymphocytes were stimulated with lipopolysaccharide (LPS, $20{\mu}g/ml$) to label $[^3H]$ thymidine. EL4 cells and lymphocytes in suspension were exposed at $0^{\circ}C$ to 0Gy, 1Gy, 5Gy, 10Gy or l5Gy for DNA single strand breaks(SSB) assay and 0Gy, 25Gy, 50Gy, 75Gy or 100Gy for DNA double strand breaks(DSB) assay of $^{60}Co$ radiation and elution procedure was performed at pH12.1 and 9.6. The number of DNA strand breaks increased with increasing doses of r rays. The strand scission factor(SSF) was estimated in each experiment (eluted volume 21ml). The slope of SSB EL4 cells was $0.01301{\pm}0.00096Gy^{-1}$ (n=5), the slope of SSB for lymphocytes was $0.01097{\pm}0.00091Gy^{-1}$ (n=5) and the slope of DSB for lymphocytes was $0.001707{\pm}0.0000573Gy^{-1}$ (n=5). Thus EL4 cells were more sensitive to induction of DNA SSB by ionizing radiation than lymphocytes (p<0.005). The ratio of slope of dose-response relationship (SSF versus dose) of lymphocytes DNA SSB as compared with the slope of DNA DSB was 6.4.

• Molecules and Cells
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• 제39권7호
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• pp.550-556
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• 2016

During meiosis, exchange of DNA segments occurs between paired homologous chromosomes in order to produce recombinant chromosomes, helping to increase genetic diversity within a species. This genetic exchange process is tightly controlled by the eukaryotic RecA homologs Rad51 and Dmc1, which are involved in strand exchange of meiotic recombination, with Rad51 participating specifically in mitotic recombination. Meiotic recombination requires an interaction between homologous chromosomes to repair programmed double-strand breaks (DSBs). In this study, we investigated the budding yeast meiosis-specific proteins Hop2 and Sae3, which function in the Dmc1-dependent pathway. This pathway mediates the homology searching and strand invasion processes. Mek1 kinase participates in switching meiotic recombination from sister bias to homolog bias after DSB formation. In the absence of Hop2 and Sae3, DSBs were produced normally, but showed defects in the DSB-to-single-end invasion transition mediated by Dmc1 and auxiliary factors, and mutant strains failed to complete proper chromosome segregation. However, in the absence of Mek1 kinase activity, Rad51-dependent recombination progressed via sister bias in the $hop2{\Delta}$ or $sae3{\Delta}$ mutants, even in the presence of Dmc1. Thus, Hop2 and Sae3 actively modulate Dmc1-dependent recombination, effectively progressing homolog bias, a process requiring Mek1 kinase activation.