• Journal of Applied Biological Chemistry
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• v.56 no.1
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• pp.37-42
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• 2013

In eukaryotes, most of the genome are transcribed, however only a small proportion of total transcripts encodes for protein, thus resulting in many of noncoding RNAs. In order to recover DNA damage including DNA double-strand breaks (DSBs) eukaryotes have evolved complex mechanisms and these are processed through coordinated mechanisms of protein sensors, transducers, and effectors including RNAs. During recent years, small RNAs have been increasingly studied and gradually considered as key regulators in various aspects of biology. Upon DNA damage, small RNAs including diRNAs (DSB induced RNA) are generated in both plant and human cell lines. Inhibition of their biogenesis has severe influence on DSB repair system.

• Journal of Veterinary Clinics
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• v.21 no.3
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• pp.253-258
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• 2004

DNA double-strand break (DSB) is a serious treat for the cells including mutations, chromosome rearrangements, and even cell death if not repaired or misrepaired. Ku heterodimer regulatory DNA binding subunits (Ku70/Ku80) bound to double strand DNA breaks are able to interact with 470-kDa DNA-dependent protein kinase catalytic subunit (DNA-PKcs), and the interaction is essential for DNA-dependent protein kinase (DNA-PK) activity. The Ku80 mutants were designed to bind Ku70 but not DNA end binding activity and the peptides were treated in breast cancer cells for co-therapy strategy to see whether the targeted inhibition of DNA-dependent protein kinase (DNA-PK) activity sensitized breast cancer cells to ionizing irradiation or chemotherapy drug to develop a treatment of breast tumors by targeting proteins involved in damage-signaling pathway and/or DNA repair. We designed domains of Ku80 mutants, 26 residues of amino acids (HN-26) as a control peptide or 38 (HNI-38) residues of amino acids which contain domains of the membrane-translocation hydrophobic signal sequence and the nuclear localization sequence, but HNI-38 has additional twelve residues of peptide inhibitor region. We observed that the synthesized peptide (HNI-38) prevented DNA-PKcs from binding to Ku70/Ku80, resulting in inactivation of DNA-PK complex activity in breast cancer cells (MDA-465 and MDA-468). Consequently, the peptide treated cells exhibited poor to no DNA repair, and became highly sensitive to irradiation or chemotherapy drugs. The growth of breast cancer cells was also inhibited. These results demonstrate the possibility of synthetic peptide to apply breast cancer therapy to induce apoptosis of cancer cells.

• Development and Reproduction
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• v.20 no.2
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• pp.141-147
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• 2016

Rad51 is a key component of homologous recombination (HR) to repair DNA double-strand breaks and it forms Rad51 recombinase filaments of broken single-stranded DNA to promote HR. In addition to its role in DNA repair and cell cycle progression, Rad51 contributes to the reprogramming process during the generation of induced pluripotent stem cells. In light of this, we performed reprogramming experiments to examine the effect of co-expression of Rad51 and four reprogramming factors, Oct4, Sox2, Klf4, and c-Myc, on the reprogramming efficiency. Co-expression of Rad51 significantly increased the numbers of alkaline phosphatase-positive colonies and embryonic stem cell-like colonies during the process of reprogramming. Co-expression ofRad51 significantly increased the expression of epithelial markers at an early stage of reprogramming compared with control cells. Phosphorylated histone H2AX (${\gamma}H2AX$), which initiates the DNA double-strand break repair system, was highly accumulated in reprogramming intermediates upon co-expression of Rad51. This study identified a novel role of Rad51 in enhancing the reprogramming efficiency, possibly by facilitating mesenchymal-to-epithelial transition and by regulating a DNA damage repair pathway during the early phase of the reprogramming process.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.10
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• pp.4879-4881
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• 2012

Epidermal growth factor receptor (EGFR) overexpression is associated with resistance to chemotherapy and radiotherapy. The EGFR modulates DNA repair after radiation-induced damage through an association with the catalytic subunit of DNA protein kinase. DNA double-strand breaks (DSBs) are the most lethal type of DNA damage induced by ionizing radiation, and non-homologous end joining is the predominant pathway for repair of radiation-induced DSBs. Some cell signaling pathways that respond to normal growth factors are abnormally activated in human cancer. These pathways also invoke the cell survival mechanisms that lead to resistance to radiation. The molecular connection between the EGFR and its control over DNA repair capacity appears to be mediated by one or more signaling pathways downstream of this receptor. The purpose of this mini-review was not only to highlight the relation of the EGFR signal as a regulatory mechanism to DNA repair and radiation resistance, but also to provide clues to improving existing radiation resistance through novel therapies based on the above-mentioned mechanism.

• Journal of Microbiology and Biotechnology
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• v.25 no.5
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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.

• BMB Reports
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• v.51 no.9
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• pp.437-443
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• 2018

Non-homologous end joining (NHEJ), and to a lesser extent, the error-free pathway known as homology-directed repair (HDR) are cellular mechanisms for recovery from double-strand DNA breaks (DSB) induced by RNA-guided programmable nuclease CRISPR/Cas. Since NHEJ is equivalent to using a duck tape to stick two pieces of metals together, the outcome of this repair mechanism is prone to error. Any out-of-frame mutations or premature stop codons resulting from NHEJ repair mechanism are extremely handy for loss-of-function studies. Substitution of a mutation on the genome with the correct exogenous repair DNA requires coordination via an error-free HDR, for targeted transgenesis. However, several practical limitations exist in harnessing the potential of HDR to replace a faulty mutation for therapeutic purposes in all cell types and more so in somatic cells. In germ cells after the DSB, copying occurs from the homologous chromosome, which increases the chances of incorporation of exogenous DNA with some degree of homology into the genome compared with somatic cells where copying from the identical sister chromatid is always preferred. This review summarizes several strategies that have been implemented to increase the frequency of HDR with a focus on somatic cells. It also highlights the limitations of this technology in gene therapy and suggests specific solutions to circumvent those barriers.

• BMB Reports
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• v.52 no.2
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• pp.151-156
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• 2019

RAD51 recombinase plays a critical role in homologous recombination and DNA damage repair. Here we showed that expression of RAD51 is frequently upregulated in lung cancer tumors compared with normal tissues and is associated with poor survival (hazard ratio (HR) = 2, P = 0.0009). Systematic investigation of lung cancer cell lines revealed higher expression of RAD51 in KRAS mutant (MT) cells compared to wildtype (WT) cells. We further showed that MT KRAS, but not WT KRAS, played a critical role in RAD51 overexpression via MYC. Moreover, our results revealed that KRAS MT cells are highly dependent on RAD51 for survival and depletion of RAD51 resulted in enhanced DNA double strand breaks, defective colony formation and cell death. Together, our results suggest that mutant KRAS promotes RAD51 expression to enhance DNA damage repair and lung cancer cell survival, suggesting that RAD51 may be an effective therapeutic target to overcome chemo/radioresistance in KRAS mutant cancers.

• BMB Reports
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• v.52 no.10
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• pp.607-612
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• 2019

During meiosis, programmed double-strand breaks (DSBs) are repaired via recombination pathways that are required for faithful chromosomal segregation and genetic diversity. In meiotic progression, the non-homologous end joining (NHEJ) pathway is suppressed and instead meiotic recombination initiated by nucleolytic resection of DSB ends is the major pathway employed. This requires diverse recombinase proteins and regulatory factors involved in the formation of crossovers (COs) and non-crossovers (NCOs). In mitosis, spontaneous DSBs occurring at the G1 phase are predominantly repaired via NHEJ, mediating the joining of DNA ends. The Ku complex binds to these DSB ends, inhibiting additional DSB resection and mediating end joining with Dnl4, Lif1, and Nej1, which join the Ku complex and DSB ends. Here, we report the role of the Ku complex in DSB repair using a physical analysis of recombination in Saccharomyces cerevisiae during meiosis. We found that the Ku complex is not essential for meiotic progression, DSB formation, joint molecule formation, or CO/NCO formation during normal meiosis. Surprisingly, in the absence of the Ku complex and functional Mre11-Rad50-Xrs2 (MRX) complex, a large portion of meiotic DSBs was repaired via the recombination pathway to form COs and NCOs. Our data suggested that Ku complex prevents meiotic recombination in the elimination of MRX activity.

• Animal cells and systems
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• v.13 no.4
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• pp.405-409
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• 2009

When DNA double-strand breaks (DSBs) were induced in mammalian cells, many DNA damage response proteins are accumulated at damage sites to form nuclear foci called IR-induced foci. Although the formation of foci has been shown to promote repair efficiency, the structural organization of chromatin in foci remains obscure. BAF53 is an actin-related protein which is required for maintenance of chromosome territory. In this study, we show that the formation of IR-induced foci by $\gamma$-H2AX and 53BP1 were reduced when BAF53 is depleted, while DSB- activated ATM pathway and the phosphorylation of H2AX remains intact after DNA damage in BAF53 knockdown cells. We also found that DSB repair efficiency was largely compromised in BAF53 knockdown cells. These results indicate that BAF53 is critical for formation of foci by $\gamma$-H2AX decorated chromatin at damage sites and the structural organization of chromatin in foci is an important factor to achieve the maximum efficiency of DNA repair.

• Journal of Plant Biotechnology
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• v.42 no.3
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• pp.154-160
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• 2015

The plant breeding technology was developed with genetic engineering. Many researchers and breeders have turned from traditional breeding to molecular breeding. Genetically modified organisms (GMO) were developed via molecular breeding technology. Currently, molecular breeding technologies facilitate efficient plant breeding without introducing foreign genes, in virtue by of gene editing technology. Gene targeting (GT) via homologous recombination (HR) is one of the best gene editing methods available to modify specific DNA sequences in genomes. GT utilizes DNA repair pathways. Thus, DNA repair systems are controlled to enhance HR processing. Engineered sequence specific endonucleases were applied to improve GT efficiency. Engineered sequence specific endonucleases like the zinc finger nuclease (ZFN), TAL effector nuclease (TALEN), and CRISPR-Cas9 create DNA double-strand breaks (DSB) that can stimulate HR at a target site. RecQl4, Exo1 and Rad51 are effectors that enhance DSB repair via the HR pathway. This review focuses on recent developments in engineered sequence specific endonucleases and ways to improve the efficiency of GT via HR effectors in plants.