• Journal of Microbiology and Biotechnology
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• v.21 no.5
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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.

• Molecules and Cells
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• v.26 no.2
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• pp.131-139
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• 2008

Expression of the seven open reading frames (ORFs) of single-stranded DNA Curtoviruses such as Beet curly top virus (BCTV) and Beet severe curly top virus (BSCTV) is driven by a bi-directional promoter. To investigate this bidirectional promoter activity with respect to viral late gene expression, transgenic Arabidopsis plants expressing a GUS reporter gene under the control of either the BCTV or BSCTV bi-directional promoter were constructed. Transgenic plants harboring constructs showed higher expression levels when the promoter of the less virulent BCTV was used than when the promoter of the more virulent BSCTV was used. In transgenic seedlings, the reporter gene constructs were expressed primarily in actively dividing tissues such as root tips and apical meristems. As the transgenic plants matured, reporter gene expression diminished but viral infection of mature transgenic plants restored reporter gene expression, particularly in transgenic plants containing BCTV virion-sense gene promoter constructs. A 30 base pair conserved late element (CLE) motif was identified that was present three times in tandem in the BCTV promoter and once in that of BSCTV. Progressive deletion of these repeats from the BCTV promoter resulted in decreased reporter gene expression, but BSCTV promoters in which one or two extra copies of this motif were inserted did not exhibit increased late gene promoter activity. These results demonstrate that Curtovirus late gene expression by virion-sense promoters depends on the developmental stage of the host plant as well as on the number of CLE motifs present in the promoter.

• Proceedings of the Zoological Society Korea Conference
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• 1998.10b
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• pp.114-114
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• 1998

No Abstract, See Full Text

• Proceedings of the Zoological Society Korea Conference
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• 1998.10b
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• pp.348.1-348
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• 1998

No Abstract, See Full Text

• Interdisciplinary Bio Central
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• v.3 no.3
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• pp.10.1-10.8
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• 2011

Introduction: Hash functions are computer algorithms that protect information and secure transactions. In response to the NIST's "International Call for Hash Function", we developed a biological hash function using the computing capabilities of bacteria. We designed a DNA-based XOR logic gate that allows bacterial colonies arranged in a series on an agar plate to perform hash function calculations. Results and Discussion: In order to provide each colony with adequate time to process inputs and perform XOR logic, we designed and successfully demonstrated a system for time-delayed bacterial growth. Our system is based on the diffusion of ${\ss}$-lactamase, resulting in destruction of ampicillin. Our DNA-based XOR logic gate design is based on the op-position of two promoters. Our results showed that $P_{lux}$ and $P_{OmpC}$ functioned as expected individually, but $P_{lux}$ did not behave as expected in the XOR construct. Our data showed that, contrary to literature reports, the $P_{lux}$ promoter is bidirectional. In the absence of the 3OC6 inducer, the LuxR activator can bind to the $P_{lux}$ promoter and induce backwards transcription. Conclusion and Prospects: Our system of time delayed bacterial growth allows for the successive processing of a bacterial hash function, and is expected to have utility in other synthetic biology applications. While testing our DNA-based XOR logic gate, we uncovered a novel function of $P_{lux}$. In the absence of autoinducer 3OC6, LuxR binds to $P_{lux}$ and activates backwards transcription. This result advances basic research and has important implications for the widespread use of the $P_{lux}$ promoter.

• Bulletin of the Korean Chemical Society
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• v.32 no.9
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• pp.3489-3492
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• 2011

• Journal of Microbiology and Biotechnology
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• v.18 no.10
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• pp.1634-1640
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• 2008

This study was focused on the screening of valuable genetic resources, such as promoters from metagenome, and describes a promoter trapping system with a bidirectional probe concept, which can select promoters or operons from various biological resources including metagenomic DNA. A pair of reporters, GFP and DsRed, facing the opposite direction without promoters, is an effective system that can function regardless of the direction of inserted promoters. The feasibility of this system was tested for the isolation of constitutively expressed promoters in E. coli from a soil metagenome, resulting in a potential pool of various promoters for practical application. The analyses of structural organization of the trapped genes demonstrated that constitutively expressible promoters in E. coli were broadly distributed within the metagenome, and suggested that some promoters were useful for the construction of expression vectors. Based on these observations, three constitutive promoters were employed in the expression vector system and their potentials for practical application were evaluated in terms of expression level, protein solubility, and effects on host growth.

• Microbiology and Biotechnology Letters
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• v.36 no.4
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• pp.260-267
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• 2008

The choice of expression vector is very important for industrial production of proteins. Therefore, the systematic mining of promoters over a wider range of genetic resource and/or host is required. We previously reported a novel bidirectional reporting system (pBGR) for the isolation of promoters from metagenome and screened useful promoters that functioned constitutively in E. coli under general culture conditions. Among them, three promoter sequences including each upstream region were amplified by PCR and used to construct new expression vectors. To facilitate subcloning, a multi-cloning site was incorporated into the downstream region of the revere primer sequence. At these sites, GFP, esterase and $\beta$-glucosidase were subcloned and analyzed the constitutive expression ability of new promoter in terms of protein solubility and expression level. As a result, these vectors expressed the proteins constitutively to a level of $2{\sim}3%$ of the total cell protein in soluble fraction (>80 %). This study suggested that excavation of metagenomic promoters for construction of expression vector in a certain strain could provide a way for the development of the expression systems.

• BMB Reports
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• v.52 no.2
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• pp.157-162
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• 2019

Our previous study found that two novel cancer-related genes, PRR11 and SKA2, constituted a classic gene pair that was regulated by p53 and NF-Y in lung cancer. However, their role and regulatory mechanism in breast cancer remain elusive. In this study, we found that the expression levels of PRR11 and SKA2 were upregulated and have a negative prognotic value in breast cancer. Loss-of-function experiments showed that RNAi-mediated knockdown of PRR11 and/or SKA2 inhibited proliferation, migration, and invasion of breast cancer cells. Mechanistic experiments revealed that knockdown of PRR11 and/or SKA2 caused dysregulation of several downstream genes, including CDK6, TPM3, and USP12, etc. Luciferase reporter assays demonstrated that wild type p53 significantly repressed the PRR11-SKA2 bidirectional promoter activity, but not NF-Y. Interestingly, NF-Y was only essential for and correlated with the expression of PRR11, but not SKA2. Consistently, adriamycin-induced (ADR) activation of endogenous p53 also caused significant repression of the PRR11 and SKA2 gene pair expression. Notably, breast cancer patients with lower expression levels of either PRR11 or SKA2, along with wild type p53, exhibited better disease-free survival compared to others with p53 mutations and/or higher expression levels of either PRR11 or SKA2. Collectively, our study indicates that the PRR11 and SKA2 transcription unit might be an oncogenic contributor and might serve as a novel diagnostic and therapeutic target in breast cancer.

• Journal of Microbiology and Biotechnology
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• v.18 no.10
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• pp.1722-1728
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• 2008

The natural course of chronic hepatitis B (CH-B) virus infection is reportedly variable, and the long-term outcomes in hepatitis B e antigen (HBeAg)-negative chronic hepatitis B infection are distinct from HBeAg-positive chronic hepatitis. However, the molecular virological factors that contribute to the progression of liver disease in the south Indian setting remain largely unclear. We prospectively studied 679 consecutive patients for HBsAg, HBeAg, anti-HBe, and HBV DNA by qualitative PCR. Randomly selected samples were subjected to bidirectional sequencing to reveal core/precore variants. Of the total 679 chronic HBV cases investigated, 23% (154/679) were replicative HBV carriers. Furthermore, amongst the 560 HBV DNA samples analyzed, 26% (146/560) were viremic. Among the 154 HBeAg positive cases, HBV DNA was positive in 118 cases (77%), significantly (p<0.001) higher than the anti-HBe positive (7%) (28/406) cases. Significant increase in liver disease (p<0.01) with ALT enzyme elevation (p<0.001) was observed in both HBe and anti-HBe viremic cases. Interestingly, low frequencies of mutations were seen in the precore region of the HBV strains studied. HBV precore and core promoter variants were less often detected in subjects with "e" negative chronic HBV infection and, therefore, may not have a prognostic role in determining liver disease sequelae in this part of tropical India.