• Journal of Microbiology and Biotechnology
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• v.19 no.3
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• pp.331-337
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• 2009

Interleukin-13 (IL-13) has been proposed as a therapeutic target for bronchial asthma as it plays crucial roles in the pathogenesis of the disease. We developed an in vitro test system measuring transcriptional downregulatory activities on IL-13 as a primary screening method to select drug candidates from natural products. The promoter region of IL-13 (-2,048 to +1) was cloned into the upstream of a luciferase gene in the plasmid pGL4.14 containing the hygromycin resistance gene as a selection marker, generating pGL4.14-IL-13. The EL-4 thymoma and RBL-2H3 mast cells transiently expressing this plasmid highly produced the luciferase activities by responding to PI (PMA and ionomycin) stimulation up to 8-fold and 13-fold compared with the control, respectively, whereas cyclosporin A, a well-known antiasthmatic agent, significantly downregulated the activities. The BF1 clone of RBL-2H3 cells constitutively expressing pGL4.14-IL-13 was established by selecting surviving cells under a constant lethal dose of hygromycin treatment. The feasibility of this system was evaluated by measuring the downregulatory activities of 354 natural products on the IL-13 promoter using the BF1 clone. An extract from Morus bombycis (named TBRC 156) significantly inhibited PI-induced luciferase activities and IL-13 mRNA expression, but not the protein expression. Fisetin (named TBRC 353) inhibited not only PI-induced luciferase activities and mRNA expression, but also the IL-13 protein secretion, whereas myricetin (named TBRC 354) could not suppress the IL-13 expression at all. Our data indicated that this in vitro test system is able to discriminate the effects on IL-13 expression, and furthermore, that it might be suitable as a simple and time-saving primary screening system to select antiasthmatic agents by measuring transcriptional activities of the IL-13 promoter.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.6
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• pp.2629-2633
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• 2012

Background: Raw betel nut (RBN) chewing is an important contributing factor for esophageal squamous cell carcinoma (ESCC), although associated genomic changes remain unclear. One difficulty in assessing the effects of exclusively RBN induced genetic alterations has been that earlier studies were performed with samples of patients commonly using tobacco and alcohol, in addition to betel-quid. Both CDKN2A (at 9p21) and Rb1 gene (at 13q14.2) are regarded as tumor suppressors involved in the development of ESCC. Therefore, the present study aimed to verify the RBN's ability to induce ESCC and assess the involvement of CDKN2A and Rb1 genes. Methods: A panel of dinucelotide polymorphic markers were chosen for loss of heterozygosity studies in 93 samples of which 34 were collected from patients with only RBN-chewing habit. Promoter hypermethylation was also investigated. Results: Loss in microsatellite markers D9S1748 and D9S1749, located close to exon $1{\beta}$ of CDKN2A/ARF gene at 9p21, was noted in 40% ESCC samples with the habit of RBN-chewing alone. Involvement of a novel site in the 9p23 region was also observed. Promoter hypermethylation of CDKN2A gene in the samples with the habit of only RBN-chewing alone was significantly higher (p=0.01) than Rb1 gene, also from the samples having the habit of use both RBN and tobacco (p=0.047). Conclusions: The data indicate that the disruption of 9p21 where CDKN2A gene resides, is the most frequent critical genetic event in RBN-associated carcinogenesis. The involvement of 9p23 as well as 13q14.2 could be required in later stages in RBN-mediated carcinogenesis.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.15
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• pp.6397-6403
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• 2014

Background: Aberrant promoter hypermethylation has been recognized in human breast carcinogenesis as a frequent molecular alteration associated with the loss of expression of a number of key regulatory genes and may serve as a biomarker. The E-cadherin gene (CDH1), mapping at chromosome 16q22, is an intercellular adhesion molecule in epithelial cells, which plays an important role in establishing and maintaining intercellular connections. The aim of our study was to assess the methylation pattern of CDH1 and to correlate it with the expression of E-cadherin, clinicopathological parameters and hormone receptor status in breast cancer patients of Kashmir. Materials and Methods: Methylation specific PCR (MSP) was used to determine the methylation status of CDH1 in 128 invasive ductal carcinomas (IDCs) paired with the corresponding normal tissue samples. Immunohistochemistry was used to study the expression of E-cadherin, ER and PR. Results: CDH1 hypermethylation was detected in 57.8% of cases and 14.8% of normal adjacent controls. Reduced levels of E-cadherin protein were observed in 71.9% of our samples. Loss of E-cadherin expression was significantly associated with the CDH1 promoter region methylation (p<0.05, OR=3.48, CI: 1.55-7.79). Hypermethylation of CDH1 was significantly associated with age at diagnosis (p=0.030), tumor size (p=0.008), tumor grade (p=0.024) and rate of node positivity or metastasis (p=0.043). Conclusions: Our preliminary findings suggest that abnormal CDH1 methylation occurs in high frequencies in infiltrating breast cancers associated with a decrease in E-cadherin expression. We found significant differences in tumor-related CDH1 gene methylation patterns relevant to tumor grade, tumor size, nodal involvement and age at diagnosis of breast tumors, which could be extended in future to provide diagnostic and prognostic information.

• BMB Reports
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• v.47 no.2
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• pp.86-91
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• 2014

Tissue-specific gene expression is regulated by epigenetic modification involving trans-acting factors. Here, we identified that the human MAGEB16 gene and its mouse homolog, Mageb16, are only expressed in the testis. To investigate the mechanism governing their expression, the promoter methylation status of these genes was examined in different samples. Two CpG islands (CGIs) in the 5' upstream region of MAGEB16 were highly demethylated in human testes, whereas they were methylated in cells without MAGEB16 expression. Similarly, the CGI in Mageb16 was hypomethylated in mouse testes but hypermethylated in other tissues and cells without Mageb16 expression. Additionally, the expression of these genes could be activated by treatment with the demethylation agent 5'-aza-2'-deoxycytidine (5'-aza-CdR). Luciferase assays revealed that both gene promoter activities were inhibited by methylation of the CGI regions. Therefore, we propose that the testis-specific expression of MAGEB16 and Mageb16 is regulated by the methylation status of their promoter regions.

• BMB Reports
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• v.49 no.10
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• pp.578-583
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• 2016

Special AT-rich sequence binding protein 1 (SATB1) is a nuclear matrix-associated DNA-binding protein that functions as a chromatin organizer. SATB1 is highly expressed in aggressive breast cancer cells and promotes growth and metastasis by reprograming gene expression. Through genome-wide cross-examination of gene expression and histone methylation, we identified SATB1 target genes for which expression is associated with altered epigenetic marks. Among the identified genes, long noncoding RNA urothelial carcinoma-associated 1 (UCA1) was upregulated by SATB1 depletion. Upregulation of UCA1 coincided with increased H3K4 trimethylation (H3K4me3) levels and decreased H3K27 trimethylation (H3K27me3) levels. Our study showed that SATB1 binds to the upstream region of UCA1 in vivo, and that its promoter activity increases with SATB1 depletion. Furthermore, simultaneous depletion of SATB1 and UCA1 potentiated suppression of tumor growth and cell survival. Thus, SATB1 repressed the expression of oncogenic UCA1, suppressing growth and survival of breast cancer cells.

• Genes and Genomics
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• v.40 no.11
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• pp.1237-1248
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• 2018

Introduction Cellulose microfibril is a major cell wall polymer that plays an important role in the growth and development of plants. The gene cellulose synthase A (CesA), encoding cellulose synthases, is involved in the synthesis of cellulose microfibrils. However, the regulatory mechanism of CesA gene expression is not well understood, especially during the early developmental stages. Objective To identify factor(s) that regulate the expression of CesA genes and ultimately control seedling growth and development. Methods The presence of cis-elements in the promoter region of the eight CesA genes identified in flax (Linum usitatissimum L. 'Nike') seedlings was verified, and three kinds of ethylene-responsive cis-elements were identified in the promoters. Therefore, the effect of ethylene on the expression of four selected CesA genes classified into Clades 1 and 6 after treatment with $10^{-4}$ and $10^{-3}M$ 1-aminocyclopropane-1-carboxylic acid (ACC) was examined in the hypocotyl of 4-6-day-old flax seedlings. Results ACC-induced ethylene either up- or down-regulated the expression of the CesA genes depending on the clade to which these genes belonged, age of seedlings, part of the hypocotyl, and concentration of ACC. Conclusion Ethylene might be one of the factors regulating the expression of CesA genes in flax seedlings.

• BMB Reports
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• v.38 no.1
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• pp.77-81
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• 2005

The monocyte chemotactic protein-3 (MCP3), on chromosome 17q11.2-q12, is a secreted chemokine, which attracts macrophages during inflammation and metastasis. In an effort to discover additional polymorphism(s) in genes whose variant(s) have been implicated in asthma, we scrutinized the genetic polymorphisms in MCP3 to evaluate it as a potential candidate gene for asthma host genetic study. By direct DNA sequencing in twenty-four individuals, we identified four sequence variants within the 3 kb full genome including 1,000bp promoter region of MCP3; one in promoter region (-420T>C), three in intron (+136C>G, +563C>T, +984G>A) respectively. The frequencies of those four SNPs were 0.020 (-420T>C), 0.038 (+136C>G), 0.080 (+563C>T), 0.035 (+984G>A), respectively, in Korean population (n = 598). Haplotypes, their frequencies and linkage disequilibrium coefficients (|D'|) between SNP pairs were estimated. The associations with the risk of asthma, skin-test reactivity and total serum IgE levels were analyzed. Using statistical analyses for association of MCP3 polymorphisms with asthma development and asthma-related phenotypes, no significant signals were detected. In conclusion, we identified four genetic polymorphisms in the important MCP3 gene, but no significant associations of MCP3 variants with asthma phenotypes were detected. MCP3 variation/haplotype information identified in this study will provide valuable information for future association studies of other allergic diseases.

• Nutrition Research and Practice
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• v.9 no.4
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• pp.379-384
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• 2015

BACKGROUND/OBJECTIVE: Apolipoprotein A5 gene promoter region T-1131C polymorphism (APOA5 T-1131C) is known to be associated with elevated plasma TG levels, although little is known of the influence of the interaction between APOA5 T-1131C and lifestyle modification on TG levels. To investigate this matter, we studied APOA5 T-1131C and plasma TG levels of subjects participating in a three-month lifestyle modification program. SUBJECTS/METHODS: A three-month lifestyle modification program was conducted with 297 participants (Age: $57{\pm}8years$) in Izumo City, Japan, from 2001-2007. Changes in energy balance (the difference between energy intake and energy expenditure) and BMI were used to evaluate the participants' responses to the lifestyle modification. RESULTS: Even after adjusting for confounding factors, plasma TG levels were significantly different at baseline among three genotype subgroups: TT, $126{\pm}68mg/dl$; TC, $134{\pm}74mg/dl$; and CC, $172{\pm}101mg/dl$. Lifestyle modification resulted in significant reductions in plasma TG levels in the TT, TC, and CC genotype subgroups: $-21.9{\pm}61.0mg/dl$, $-20.9{\pm}51.0mg/dl$, and $-42.6{\pm}78.5mg/dl$, respectively, with no significant differences between them. In a stepwise regression analysis, age, APOA5 T-1131C, body mass index (BMI), homeostasis model assessment-insulin resistance (HOMA-IR), and the 18:1/18:0 ratio showed independent association with plasma TG levels at baseline. In a general linear model analysis, APOA5 T-1131C C-allele carriers showed significantly greater TG reduction with decreased energy balance than wild type carriers after adjustment for age, gender, and baseline plasma TG levels. CONCLUSIONS: The genetic effects of APOA5 T-1131C independently affected plasma TG levels. However, lifestyle modification was effective in significantly reducing plasma TG levels despite the APOA5 T-1131C genotype background.

• The Journal of the Korean Society for Microbiology
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• v.35 no.3
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• pp.263-271
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• 2000

A clinical isolate of Klebsiella pneumoniae K7746 produced the extended-spectrum ${\beta}$-lactamase (ESBL) SHV-12. A 6.6 kb BamHI fragment containing the $bla_{SHV-12}$ gene of K7746 strain was cloned into pCRScriptCAM vector resulting in the recombinant plasmid p7746-Cl. The restriction map of 3.6 kb inserted DNA and sequences immediately surrounding $bla_{SHV-12}$ of p7746-C1 were homologous to plasmid pMPA2a carrying $bla_{SHV-2a}$. In addition, both $bla_{SHV-12}$ and $bla_{SHV-2a}$ were expressed from a common hybrid promoter made of the -35 region derived from the left inverted repeat of IS26 and the -10 region from the $bla_{SHV}$ promoter itself. The results indicate that $bla_{SHV-12}$ and $bla_{SHV-2a}$ may have evolved from a common ancestor in the sequential order of $bla_{SHV-2a}$ first, followed by $bla_{SHV-12}$. Furthermore, by the PCR mapping method using primers corresponding to the IS26 and $bla_{SHV}$, the association between IS26 and $bla_{SHV}$ was studied in 12 clinical isolates carrying $bla_{SHV-2a}$, 27 clinical isolates carrying $bla_{SHV-12}$, and 5 reference strains carrying $bla_{SHV-1}$ to $bla_{SHV-5}$. All 39 strains carrying $bla_{SHV-2a}$ or $bla_{SHV-12}$ were positive by the PCR, providing confirmative evidence that IS26 has been involved in the evolution and dissemination of $bla_{SHV-2a}$ and $bla_{SHV-12}$. But 5 reference strains carrying $bla_{SHV-1}$ to $bla_{SHV-5}$ were negative by the PCR. Therefore, we concluded that the molecular evolutionary pathway of $bla_{SHV-2a}$ and $bla_{SHV-12}$ may be different from that of other $bla_{SHV-ESBL}$, e.g., $bla_{SHV-2}$, $bla_{SHV-3}$, $bla_{SHV-4}$, and $bla_{SHV-5}$.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.4
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• pp.2093-2097
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• 2016

Background: HCV is a major global health problem. IL-27 is a member of the IL-6/IL-12 cytokine family with a broad range of anti-inflammatory properties. Recent studies highlighted the effect of a SNP in the IL-27 promoter region on modulating the progression of infectious diseases and individual responses to therapy. Aim of the work: The present study investigated the potential role of (-964 A/G) SNP in the promoter region of IL-27p28 gene (alleles rs153109) on the outcome of HCV infection among genotype 4a infected patients. Materials and Methods: HCV genotyping confirmed that all of the HCV-infected patients had genotype 4a infection. Genomic DNA was extracted from 111 patients with chronic HCV infection, 42 spontaneous resolvers (SR) and 16 healthy controls. IL- 27p28.rs153109 genotyping was assessed using PCR-RFLP then confirmed by DNA sequencing. Results: The frequency of IL-27-p28.rs153109AA, AG, and GG genotypes among chronically infected subjects were 74.8 %, 25.2%, and 0% while among the SR, they were 57.1%, 35.7%, and 7.14%, respectively. Our data show the unique presence of G/G genotype in the SR group (3 patients; 7.14%). Moreover, the "G" allele frequencies among chronic and resolved subjects were 12.6% and 25.0%, respectively (p=0.0136). Importantly, subjects with the GG genotype were more likely to clear their HCV infection than those with the AA genotype (p=0.0118). Conclusions: HCV genotype 4a subjects with the IL-27-p28.rs153109 A/G and G/G genotype were more likely to clear their HCV infection. Therefore, we propose IL- 27p28.rs153109SNPas a genetic biomarker for predicting HCV infection outcome.