• Journal of Microbiology and Biotechnology
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• 제21권5호
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• pp.455-463
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• 2011

Metallothioneins are a class of small cysteine-rich proteins that have been associated with increased tolerance to metal and oxidative stresses in animals, plants, and fungi. We investigated a metallothionein-like (mt-like) gene shown previously to be upregulated in fruiting bodies of the fungus Agaricus bisporus in response to post-harvest storage. Analysis of an A. bisporus genomic DNA cosmid library identified two similar mt-like genes (met1 and met2) arranged as a bidirectional gene pair transcribed from the same promoter region. The promoter contained regulatory elements including 9 metal responsive elements and a CAAT box region 220 bp downstream of met1 that showed striking similarity to a feature in Coprinopsis cinerea mt-like gene promoters. Transcriptional analysis showed that both met genes are significantly and rapidly (within 3 hours) upregulated during post-harvest storage and expression is significantly greater in stipe and cap tissues compared with the gills. However, a strong directionality of the promoter was demonstrated, as transcript levels of met1 were at least two orders of magnitude greater than those of met2 in all samples tested.

• 한국양식학회지
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• 제10권1호
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• pp.33-37
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• 1997

미꾸라지를 대상으로 어류 유전자 이식을 위한 유용 model system으로 개발하기 위한 연구의 일환으로 잉어의$\beta$-actin 유전자의 promoter와 CAT 유전자가 융합되어 있는 외래 유전자 (pFV4CAT)를 미꾸라지에 이식, transgenic founder 미꾸라지를 생산하였다. PCR 분석결과, 이식된 외래 유전자는 부화 후 9개월된 성어에서 7.4-37.0%의 빈도로 존재하였으며 Southern blot 분석에 의해서 염색체 상의 삽입 가능성을 나타내었다. In situ immunohistochemical analysis 결과 이식된 외래 유전자는 미꾸라지 세포내에서 비교적 높은 빈도로 발현하였으며 그 발현 양상은 개체마다 다양하게 나타났다.

• Journal of Microbiology
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• 제35권4호
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• pp.277-283
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• 1997

Promoters inducible by paraquat, a superocide-generating agent, were isolated from Escherichia coli using a promoter-probing plasmid pRS415 with promoterless lacA gene. Twenty one promoters induced by paraquat were selected and further characterized. From sequence analysis, thirteen of the promoters were mapped to their specific loci on the Escherichia coli chromosome. Several promoters were mapped to the upstream of known genes such as usgl, katG, and mglB, whose relationships with superoxide response have not been previously reported. Other promoters were mapped to the upstream region of unknown open reading frames. Downstream of HC 96 promoter are uncharacterized ORFs whose sequences are homologous to ABC-transporter subunits. Downstream of HC84 promoter is an ORF encoding a transcriptional regulator-like protein, which contains a LysR family-specific HTH (helix-turn-helix) DNA bindign motif. We investigated whether these promoters belong to the soxRS regulon. All promoters except HC96 were found to belong to the soxRS regulon. The HC96 promoter was significantly induced by paraquat in the soxRS deletion mutant strain. The basal transcription level of three promoters (HE43, HC71, HD94) significantly increased at the stationary phase, implying that they are regulated by RpoS. However, paraquat inducibility of all promoters disappeared in the stationary phase, suggesting that SoxRS regulatory system is active only in rapidly growing cells.

• Molecules and Cells
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• 제24권1호
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• pp.148-151
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• 2007

Porcine endogenous retroviruses (PERVs) in the pig genome represent a potential risk of infection in pig-to-human transplantation and are transmitted vertically. The solitary long terminal repeat (LTR) elements of the PERVs affect the replication properties of the individual viruses via their repeat sequences and by encoding a set of specific transcription factors. We examined the promoter activities of solitary LTR elements belonging to the PERV-A and -B families of the Korean domestic pig (KDP) using luciferase reporters. Three of the LTR structures (of PERV-A5-KDP, PERV-A7-KDP, PERV-A8-KDP) had different promoter activities in human HCT116 cells and monkey Cos7 cells, and potential negatively and positively acting regions affecting transcription were identified by deletion analysis. These data suggest that specific sequences in the U3 region of a given LTR element can affect the activities of promoter or enhancer elements in the PERV.

• Journal of Plant Biology
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• 제39권4호
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• pp.279-286
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• 1996

To know the changes of rbcL mRNA level by illumination, Northern hybridization analysis was performed with maize (Zea mays L.cv. Golden X Bantam). The average level of rbcL. mRNA in the light-grown shoots was 3.1 times higher than that of the dark-grown shoots after 6 to 10 growth days. The maximum difference of rbcL mRNA level between the dark-grown and the light-grown shoots was 5.1 folds. These results indicate that accumulation of rbcL mRNAin maize shoots is induced by light. Since the transcriptional DNA binding proteins and their cognate promoter elements, we carried out gel-retardation assays to elucidate the specific binding proteins on the rbcL promoter. It was found that plastid proteins of light-grown shoots bound to the R2 DNA fragment (-33 to -229) and R3 DNA fragment (-230 to -418 from ATG) of the rbcL promoter. From the results of competitive binding assays and heat or protease treatments, it was demonstrated that the bindings were sequence-specific DNA-protein interactions. Therefore, it could be concluded that the rbcL promoter region has at least two specific recognition sites for plastid proteins.

• BMB Reports
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• 제55권11호
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• pp.553-558
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• 2022

Hepatocellular carcinoma (HCC) is dangerous cancer that often evades early detection because it is asymptomatic and an effective detection method is lacking. For people with chronic liver inflammation who are at high risk of developing HCC, a sensitive detection method for HCC is needed. In a meta-analysis of The Cancer Genome Atlas pan-cancer methylation database, we identified a CpG island in the USP44 promoter that is methylated specifically in HCC. We developed methylation-sensitive high-resolution melting (MS-HRM) analysis to measure the methylation levels of the USP promoter in cell-free DNA isolated from patients. Our MS-HRM assay correctly identified 40% of patients with early-stage HCC, whereas the α-fetoprotein test, which is currently used to detect HCC, correctly identified only 25% of early-stage HCC patients. These results demonstrate that USP44 MS-HRM analysis is suitable for HCC surveillance.

• Asian Pacific Journal of Cancer Prevention
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• 제14권6호
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• pp.3773-3778
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• 2013

Increased oxidative stress and changes in DNA methylation are frequently detected in bladder cancer patients. We previously demonstrated a relationship between increased oxidative stress and hypomethylation of the transposable long-interspersed nuclear element-1 (LINE-1). Promoter hypermethylation of a tumor suppressor gene, runt-related transcription factor 3 (RUNX3), may also be associated with bladder cancer genesis. In this study, we investigated changes of DNA methylation in LINE-1 and RUNX3 promoter in a bladder cancer cell (UM-UC-3) under oxidative stress conditions, stimulated by challenge with $H_2O_2$ for 72 h. Cells were pretreated with an antioxidant, tocopheryl acetate for 1 h to attenuate oxidative stress. Methylation levels of LINE-1 and RUNX3 promoter were measured by combined bisulfite restriction analysis PCR and methylation-specific PCR, respectively. Levels of LINE-1 methylation were significantly decreased in $H_2O_2$-treated cells, and reestablished after pretreated with tocopheryl acetate. Methylation of RUNX3 promoter was significantly increased in cells exposed to $H_2O_2$. In tocopheryl acetate pretreated cells, it was markedly decreased. In conclusion, hypomethylation of LINE-1 and hypermethylation of RUNX3 promoter in bladder cancer cell line was experimentally induced by reactive oxygen species (ROS). The present findings support the hypothesis that oxidative stress promotes urothelial cell carcinogenesis through modulation of DNA methylation. Our data also imply that mechanistic pathways of ROS-induced alteration of DNA methylation in a repetitive DNA element and a gene promoter might differ.

• Journal of Plant Biotechnology
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• 제47권2호
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• pp.172-178
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• 2020

Fruit-specific promoters play an important role in the improvement of traits, such as fruit quality through genetic engineering. In tomato, the development of fruit-specific promoters was previously reported, but less attention has been paid to the promoters involved in the fruit development stage. In this study, we characterized the gene expression patterns of tomato alcohol dehydrogenase 2 (SlAdh2) in various tissues of wild-type tomato (cv. Ailsa Craig). Our findings revealed that SlAdh2 expression levels were higher in the developing fruit than in the leaves, stems, and flowers. The ProSlAdh2 region, which is expressed at different stages of fruit development, was isolated from tomato genomic DNA. Following this, it was fused with a β-glucuronidase reporter gene (GUS) and introduced into wild-type tomato using Agrobacterium-mediated transformation to evaluate promoter activity in the various tissues of transgenic tomato. The ProSlAdh2:GUS promoter exhibited strong activity in the fruit and weak activity in the stems, but displayed undetectable activity in the leaves and flowers. Interestingly, the promoter was active from the appearance of the green fruit (1 cm in size) to the well-ripened stage in transgenic tomatoes, indicating its suitability for transgene expression during fruit development and ripening. Thus, our findings suggest that ProSlAdh2 may serve as a potential fruit-specific promoter for genetic-based improvement of tomato fruit quality.

• Asian Pacific Journal of Cancer Prevention
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• 제13권5호
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• pp.1833-1836
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• 2012

Objective: Promoter methylation, which can be regulated by MTHFR activity, is associated with silencing of genes. In this study we evaluated the methylation status (type) of the BRCA2 promoter in ovarian cancer patients carrying different genotypes of the MTHFR gene (A or C polymorphisms at position 1298). Methods: The methylation type of the BRCA2 promoter was evaluated using bisulfate-modified DNA in methylation-specific PCR and the MTHFRa1278c polymorphism was assessed by PCR-RFLP. Results: Analysis of the BRCA2 promoter methylation type of cases showed that 7 out of 60 cases (11.7%) were methylated while the remaining 53 (88.3%) were unmethylated. In methylated cases, one out of the 7 cases had a CC genotype and the remaining 6 methylated cases had an AC genotype. The AA genotype was absent. In unmethylated cases, 34, 18, and one out of these had AC, AA and CC genotype, respectively. Conclusion: There was no significant relationship between the methylation types of the BRCA2 promoter in different genotypes of MTHFRa1298c polymorphism in ovarian cancer; p=0.255. There was no significant relation between the methylation types of the BRCA2 promoter in different genotypes of the MTHFRa1298c polymorphism in ovarian cancer.

• 미생물학회지
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• 제52권3호
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• pp.249-253
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• 2016

먹물버섯의 하나인 Coprinellus congregatus는 생활사 동안 여러 종의 laccase 효소를 생성한다. 균사 끝 효소와 버섯시원체 효소 및 sclerotium (균핵) 효소들은 모두 이 균의 분화와 관련되었다. 이핵체 균사를 산성 액체배지(pH 4.0-4.5)에 접종하면 새로운 laccase가 합성되어 분비된다. 이 laccase 유전자의 프로모터의 어느 부분이 산 충격의 신호에 관련된 단백질이 결합하는가 분석하기 위하여 녹색형광단백질(green fluorescent protein, GFP) 유전자를 laccase 프로모터 2.0 kb 다음에 연결하고, 이를 형질전환 벡터인 pBARGEM7-1에 삽입함으로써 발현벡터를 구축하였다. 이 promoter-GFP 조합의 5'-region부터 차례로 제거한 짧은 길이의 이 발현벡터를 먹물버섯 교배형 a1균과 a2균에 형질전환 방법으로 도입시키고 phosphinothricin 저항성으로 형질전환체들을 선발하였다. 선발된 형질전환체 a1 (a1TF)과 a2 (a2TF)를 서로 교배하여 동형접합(homozygotic) 이핵체 형질전환체를 만들었다. 이들을 산성 액체배지에서 36시간 배양하고 균체를 모아 confocal microscope를 사용하여 형광을 분석하였다. Laccase 유전자의 전체 프로모터(2.0 kb)를 가진 발현벡터(F0-GFP)를 도입한 동형접합 형질전환체에서는 형광을 보였으나, 그 보다 짧은 길이(1.29 kb 이하)의 프로모터를 가진 형질전환체에서는 형광이 나타나지 않았다. 이 결과에 근거하여 먹물버섯의 산 충격에 대한 신호를 받는 부위가 laccase 유전자 프로모터의 -2.0 kb ~ -1.29 kb 사이에 있을 것으로 추정한다.