• Asian Pacific Journal of Cancer Prevention
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• v.14 no.1
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• pp.359-365
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• 2013

The high mobility group box 1 (HMGB1) protein is a multifunctional cytokine-like molecule that plays an important role in the pathogenesis of tumors. In this study, real-time polymerase chain reactions and Western blot assays indicated that HMGB1 transcriptional activity and protein level are increased in $Tax^+$-T cells (TaxP). To clarify the mechanisms, a series of HMGB1 deletion reporter plasmids (pHLuc1 to pHLuc6) were transfected into $Tax^-$-T cells (TaxN, Jurkat) and $Tax^+$-T cells (TaxP). We found that promoter activity in $Tax^+$-T cells to be higher than that in $Tax^-$-T cells, indicating a significant increase in pHLuc6. Bay11-7082 (NF-${\kappa}B$ inhibitor) treatment did not block the enhancing effect. Chromatin immunoprecipitation assays revealed that Tax was retained on a HMGB1 promoter fragment encompassing -1163 to -975. Bioinformatics analysis showed six characteristic cis-elements for CdxA, AP-1, AML-1a, USF, v-Myb, and C/EBP in the fragment in question. Mutation of cis-elements for C/EBP reduced significant HMGB1 promoter activity induced by Tax. These findings indicate that Tax enhances the expression of HMGB1 gene at the transcriptional level, possibly by interacting with C/EBP.

• Toxicological Research
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• v.18 no.2
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• pp.129-138
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• 2002

The mouse lymphoma assay (MLA) has been recently validated as a sensitive and specific test system to determine the genotoxic potential for a chemical. The objective of this study is to evaluate the utility of MLA for detecting mutagens. Especially, to compare MLA with the in vitro chromosomal aberration test (CA), we performed MLA using the microwell method with three chemicals (hydroxyurea, theophylline and amino acid copper complex), which were reportedly positive in the CA. In cell treated with hydroxyurea, anti-neoplastic agent that blocks DNA replication, evidence of a positive response was obtained without S9 mix for 4 h and 24 h. In addition, analysis of colony size distribution at concentration that gave an elevated mutant fraction showed that hydroxyurea induced a high proportion of small type colonies, indicating that hydroxyurea-induced mutation is associated with large chromosomal deletion. Conversely, negative MLA result was obtained for theophylline, which was wed as central nervous system stimulator. Although theophylline increased the mutant frequency at concentration of 1250 $\mu\textrm{g}$/$\textrm{m}{\ell}$ with S9 mix for 4 h, a concentration-related increase in mutant frequency was not observed. The MLA result of amino acid copper complex was considered equivocal because the positive result was obtained at concentration showing 10% or less RS or RTG. Thus, among 3 CA-positive chemicals, positive MLA result was obtained for one. The other two chemicals were negative and equivocal. However MLA, which evaluates mutagenic potential of chemicals through colony formation by cell grouth, may provide a higher predictivity of carcinogenesis than CA.

• Journal of the Institute of Electronics and Information Engineers
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• v.49 no.10
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• pp.91-101
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• 2012

DNA watermarking have been interested for both the security of private genetic information or huge DNA storage information and the copyright protection of GMO. Multimedia watermarking has been mainly designed on the basis of frequency domain, such as DCT, DWT, FMT, and so on, for the robustness and invisibility. But a frequency domain watermarking for coding DNA sequence has a considerable constraint for embedding the watermark because transform and inverse transform must be performed without completely changing the amino acid sequence. This paper presents a coding sequence watermarking on lifting based DWT domain and brings up the availability of frequency domain watermarking for DNA sequence. From experimental results, we verified that the proposed scheme has the robustness to until a combination of 10% point mutations, 5% insertion and deletion mutations and also the amino preservation and the security.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.5
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• pp.743-748
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• 2017

Objective: Based on rapid advancement of genetic modification techniques, genomic editing is expected to become the most efficient tool for improvement of economic traits in livestock as well as poultry. In this study, we examined and verified the nickase of mutated CRISPR-associated protein 9 (Cas9) to modulate the specific target gene in chicken DF1 cells. Methods: Chicken myostatin which inhibits muscle cell growth and differentiation during myogenesis was targeted to be deleted and mutated by the Cas9-D10A nickase. After co-transfection of the nickase expression vector with green fluorescent gene (GFP) gene and targeted multiplex guide RNAs (gRNAs), the GFP-positive cells were sorted out by fluorescence-activated cell sorting procedure. Results: Through the genotyping analysis of the knockout cells, the mutant induction efficiency was 100% in the targeted site. Number of the deleted nucleotides ranged from 2 to 39 nucleotide deletion. There was no phenotypic difference between regular cells and knockout cells. However, myostatin protein was not apparently detected in the knockout cells by Western blotting. Additionally, six off-target sites were predicted and analyzed but any non-specific mutation in the off-target sites was not observed. Conclusion: The knockout technical platform with the nickase and multiplex gRNAs can be efficiently and stablely applied to functional genomics study in poultry and finally adapted to generate the knockout poultry for agribio industry.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.1
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• pp.253-258
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• 2015

Background: Development of squamous cell cancer of head and neck (SCCHN) is associated with human papillomavirus (HPV) infection, which in turn is closely related with expression of $p16^{INK4A}$. Loss of $p16^{INK4A}$ expression by deletion, mutation, or hypermethylation is common in SCCHN. We here evaluated $p16^{INK4A}$ as a prognostic marker of treatment response and survival in our SCCHN patients with laryngeal, hypopharyngeal or nasopharyngeal cancers. Materials and Methods: 131 patients diagnosed with SCCHN between January 2,2006 and July 17, 2010 were examined for $p16^{INK4A}$. The median age was 60 years (15-82 years). Fifty one patients were stage I-II and 80 were stage III-IV. Immunohistochemical expression of $p16^{INK4A}$ was analyzed in pretreatment paraffin-embedded tumor blocks. The influence of $p16^{INK4A}$ status on disease-free survival, and overall survival after treatment was evaluated. Results: $p16^{INK4A}$ positivity was found in 58 patients (44%). Tumor-positivity for$ p16^{INK4A}$ was correlated with improved disease free survival (70.1 months vs 59 months) and improved overall survival (2, 3 and 5-year values; 77% vs 72%, 70% vs 63% and, 63% vs 55%; respectively). On multivariate analysis, stage was determined as independent prognostic factor for disease-free survival. Conclusions: Stage was the major prognostic factor on treatment response and survival in our patients. $p16^{INK4A}$ status predicts better outcome in laryngeal, hypopharyngeal or nasopharyngeal cancer cases treated with surgery plus adjuvant radiochemotherapy as well as with definitive radiation therapy and/or chemotherapy.

• Development and Reproduction
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• v.15 no.1
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• pp.25-30
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• 2011

Embryonic stem (ES) cells can self-renew maintaining the undifferentiated state. Self renewal requires many factors such as Oct4, Sox2, FoxD3, and Nanog. NF-${\kappa}B$ is a transcription factor involved in many biological activities. Expression and activity of NF-${\kappa}B$ increase upon differentiation of ES cells. Reportedly, Nanog protein directly binds to NF-${\kappa}B$ protein and inhibits its activity in ES cells. Here, we found a potential binding site of NF-${\kappa}B$ in the human Nanog promoter and postulated that NF-${\kappa}B$ protein may regulate expression of the Nanog gene. We used human embryonic carcinoma (EC) cells as a model system of ES cells and confirmed decrease of Nanog and increase of NF-${\kappa}B$ upon differentiation induced by retinoic acid. Although deletion analysis on the DNA fragment including NF-${\kappa}B$ binding site suggested involvement of NF-${\kappa}B$ in the negative regulation of the promoter, site-directed mutation of NF-${\kappa}B$ binding site had no effect on the Nanog promoter activity. Furthermore, no direct association of NF-${\kappa}B$ with the Nanog promoter was detected during differentiation. Therefore, we conclude that NF-${\kappa}B$ protein may not be involved in transcriptional regulation of Nanog gene expression in EC cells and possibly in ES cells.

• Korean Journal of Microbiology
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• v.37 no.1
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• pp.42-48
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• 2001

Allele rescue and sequence analysis of soo1-1 allele in Saccharomyces cerevisiae mutant LP0353 revealed that soo1-1 is identical to the previously reported ret1-1 allele, which has a base substitution of A for $G^{681}$ leading to an amino acid substitution of aspartic acid for $glycine^{227}$ in Soolp. However, it was revealed that the addition of osmotic stabilizer, such as 1.2M sorbitol can rescue the temperature sensitive phenotype of the ret1-1 mutant and that the soo1-1/ret1-1 mutation may confer defects in post-translational modification of proteins involved in the yeast cell wall biogenesis. Evidence for a putative role of 5th WD40 domain of the Soo1p/$\alpha$-COP in the construction and maintenance of cell walls was also presented by complementation test with deletion constructs of the SOOl.

• Korean Journal of Environmental Biology
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• v.20 no.4
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• pp.316-324
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• 2002

To investigate the mutational spectrum of laccase (Lac) genes (lac -A and lac -B) involved in degrading lignin which is the recalcitrant cell wall polymer, the genes of the Pleurotus ostreatus mutants induced by gamma ray radiation were amplified by PCR and were cloned. All partial lac-A genes of 4 mutants (PO-6, -7, -14 and -15) consisted of 1763 base pairs due to the deletion of two bases (491-nt and 492-nt) and addition of one base (875-nt) in 1764 base pairs of lac -A gene of PO-1. Totally 36 mutational hot spots were detected and 32 positions were mutated in all of those 4 mutants simultaneously. These mutations were predominantly A : T -> G : C transitions (40%). Putative amino acid sequences of lac -A genes of mutants have one simultaneous mutated residue (from Thr-44 to Ala-44). The 1764 bp of partial lac -B gene was cloned only in PO -5 mutant and contained 19 mutated bases. These mutations were predominantly G : C->A : T transitions (45%). Lac-B protein of PO-5 has two mutated residues of Glu-290 and His-363 from Ala-290 and Phe-363, respectively. The hyper-mutational positions were concentrated in specific regions of between 50-nt and 900-nt in lac genes. These results suggested that the mutational hotspots responded to gamma radiation could be in some genes, at least lac -A and lac -B of p. ostreatus.

• Molecules and Cells
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• v.22 no.2
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• pp.146-153
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• 2006

Cell polarity is critical for the division, differentiation, migration, and signaling of eukaryotic cells. RAX2 of budding yeast encodes a membrane protein localized at the cell cortex that helps maintain the polarity of the bipolar pattern. Here, we designate SPAC6f6.06c as $rax2^+$ of Schizosaccharomyces pombe, based on its sequence homology with RAX2, and examine its function in cell polarity. S. pombe $rax2^+$ is not essential, but ${\Delta}rax2$ cells are slightly smaller and grow slower than wild type cells. During vegetative growth or arrest at G1 by mutation of cdc10, deletion of $rax2^+$ increases the number of cells failing old end growth just after division. In addition, this failure of old end growth is dramatically increased in ${\Delta}tea1{\Delta}rax2$, pointing to genetic interaction of $rax2^+$ with $tea1^+$. ${\Delta}rax2$ cells contain normal actin and microtubule cytoskeletons, but lack actin cables, and the polarity factor for3p is not properly localized at the growing tip. In ${\Delta}rax2$ cells, and endogenous rax2p is localized at the cell cortex of growing cell tips in an actin- and microtubule-dependent manner. However, ${\Delta}rax2$ cells show no defects in cell polarity during shmoo formation and conjugation. Taken together, these observations suggest that rax2p controls the cell polarity of fission yeast during vegetative growth by regulating for3p localization.

• Korean Journal of Medicinal Crop Science
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• v.3 no.1
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• pp.50-55
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• 1995

This study was conducted to determine the optimum conditions for induction and different somatic of somatic embryos as well as germination of encapsulated and stored somatic embryos. Somatic embryos was better formed in 1/2X MS medium than full - strength MS medium. 0.1 to 1.0mg/lBA and kinetin promoted shoot differentiation of somatic embryos. Higher concentration tend to inhibit differentiation. IAA affect positively both root and shoot growth. In vitro germination of somatic embryos encapsulated with 2% alginate matrix containing 1/2 MS nutrient medium and $AgNO_3$ 5mg/l was 86%. Storage of somatic embryos was effecive at $5^{\circ}C$ but the germination rate decreased with longer storage period. RAPD analyses with plants regenerated from the somatic embryos showed DNA polymorphism, indicating abolition of primer binding site by point mutation, deletion, or insertion of certain sequences.