• 대한의생명과학회지
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• 제23권3호
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• pp.272-276
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• 2017

HOX genes are transcription factors that play important roles in body patterning and cell fate specification during normal development. In previous study, we found aberrant overexpression of HOXB5 in breast cancer tissues and cell lines, and demonstrated that HOXB5 is important in regulation of cell proliferation, tamoxifen resistance, and invasiveness through the epithelial-mesenchymal transition (EMT). Although the relationship between HOXB5 and phenotypic changes in MCF7 breast cancer cells has been studied, the molecular function of HOXB5 as a transcription factor remains unclear. IL-6 has been reported to be involved in not only inflammation but also cancer progression, which is characterized by the increase of growth speed and invasiveness of tumor cells. In this study, we selected Interleukin-6 (IL-6) as HOXB5 putative downstream target gene and discovered that HOXB5 transcriptionally up-regulated the expression of IL-6 in HOXB5 overexpressing MCF7 cells. The upstream region (~1.2 kb) of IL-6 promoter turned out to contain several putative HOX consensus binding sites. Chromatin immunoprecipitation assay confirmed that HOXB5 directly binds to the promoter region of IL-6 and positively regulated the expression of IL-6. These data all together, indicate that HOXB5 promotes IL-6 transcription by actively binding to the putative binding sites located in the upstream region of IL-6, which enable to increase its promoter activity in MCF7 breast cancer cells.

• 대한의생명과학회지
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• 제12권3호
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• pp.139-143
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• 2006

Jopock (Jpk) has previously been ascertained that induces both bacterial and mammalian cell death. The Escherichia coli cells expressing Glutathion S-transferase (GST) fused Jpk showed elongated phenotype and inhibited cell growth which led eventual cell death. In an attempt to search the genetic suppressor of the lethal protein Jpk in bacterial cells, we constructed a unidirectional protein expression vector inserting tac promoter next to the C-terminus Jpk in pGEX-Jpk. The function of additional tac promoter was confirmed by substituting lac promoter in Plac-TOPO plasmid. The cells harboring plac- TOPO, which regulates $lacZ{\alpha}$ gene expression under lac promoter, formed blue colonies in 5-bromo-4-3 $indolyo-{\beta}-D-galactoside$ (X-gal) plate. When lac promoter was changed to tac promoter, same results were observed. Since the addition of tac promoter did not affect the toxic effect of Jpk, the pGEX-Jpk-ptac could be a useful vector for the screening of suppressor(s) for Jpk, in which GST-Jpk and a putative Jpk-suppressing protein are coexpressing from two unidirectional tac promoters, which response to the same inducer, $isopropyl-{\beta}-D-thiogalactopyranoside (IPTG)$.

• Molecules and Cells
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• 제26권3호
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• pp.236-242
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• 2008

Invertase (${\beta}$-D-fructofuranosidase; EC 3.2.1.26) catalyzes the conversion of sucrose into glucose and fructose and is involved in an array of important processes, including phloem unloading, carbon partitioning, the response to pathogens, and the control of cell differentiation and development. Its importance may have caused the invertases to evolve into a multigene family whose members are regulated by a variety of different mechanisms, such as pH, sucrose levels, and inhibitor proteins. Although putative invertase inhibitors in the Arabidopsis genome are easy to locate, few studies have been conducted to elucidate their individual functions in vivo in plant growth and development because of their high redundancy. In this study we assessed the functional role of the putative invertase inhibitors in Arabidopsis by generating transgenic plants harboring a putative invertase inhibitor gene under the control of the CaMV35S promoter. A transgenic plant that expressed high levels of the putative invertase inhibitor transcript when grown under normal conditions was chosen for the current study. To our surprise, the stability of the invertase inhibitor transcripts was shown to be down-regulated by the phytohormone ABA (abscisic acid). It is well established that ABA enhances invertase activity in vivo but the underlying mechanisms are still poorly understood. Our results thus suggest that one way ABA regulates invertase activity is by down-regulating its inhibitor.

• Molecules and Cells
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• 제23권1호
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• pp.80-87
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• 2007

Beet curly top virus (BCTV) and Beet severe curly top virus (BSCTV), members of curtoviruses, encode seven open reading frames (ORFs) within a ~3 kb genome. One of these viral ORFs, C1, is known to play an important role in the early stage of viral infection in plants during initiation of viral DNA replication. We used promoter:: reporter (${\beta}$-glucuronidase) gene fusions in transgenic Arabidopsis to identify the putative promoter region of BCTV ORF C1. Unlike other geminiviruses, the intergenic region of BCTV was not sufficient to promote C1 expression in transgenic plants. When sequences extending into the coding region of C1 were tested, strong expression of the reporter protein was observed in vascular tissues of transgenic plants. This expression was not dependent on the presence of the intergenic regions or proximal 5' portions of the C1 coding region. Transgenic plants expressing a reporter gene under control of the putative complete C1 promoter were inoculated with virus to determine if any viral transcript affected C1 expression. Virus inoculated plants did not show any altered pattern or change in of reporter gene expression level. These results suggest that (1) important transcriptional activator elements for C1 expression reside in the 3' portion of C1 coding area itself, (2) C1 protein does not auto-regulate its own expression and (3) C1 expression of two curtoviruses is controlled differently compared to other geminiviruses.

• Journal of Microbiology and Biotechnology
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• 제27권9호
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• pp.1586-1592
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• 2017

Some promoters were isolated and characterized from the genome of Leuconostoc mesenteroides SY2, an isolate from kimchi, a Korean traditional fermented vegetable. Chromosomal DNA of L. mesenteroides SY2 was digested with Sau3AI and ligated with BamHI-cut pBV5030, a promoter screening vector containing a promoterless cat-86. Among E. coli transformants (TFs) resistant against Cm (chloramphenicol), 17 were able to grow in the presence of $1,000{\mu}g/ml$ Cm and their inserts were sequenced. Transcription start sites were examined for three putative promoters (P04C, P25C, and P33C) by primer extension. Four putative promoters were inserted upstream of a promoterless ${\alpha}$-amylase reporter gene in $pJY15{\alpha}$. ${\alpha}$-Amylase activities of E. coli TFs containing $pJY15{\alpha}$ (control, no promoter), $pJY03{\alpha}$ ($pJY15{\alpha}$ with P03C), $pJY04{\alpha}$ (with P04C), $pJY25{\alpha}$ (with P25C), and $pJY33{\alpha}$ (with P33C) were 66.9, 78.7, 122.1, 70.8, and 99.3 U, respectively. Cells harboring $pJY04{\alpha}$ showed 1.8 times higher activity than the control. Some promoters characterized in this study might be useful for construction of food-grade expression vectors for Leuconostoc sp. and related lactic acid bacteria.

• Journal of Microbiology and Biotechnology
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• 제1권4호
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• pp.240-245
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• 1991

We have constructed a promoter-probe vector pKU20 using pKT230, a derivative of broad-host-range plsmid RSF1010, as a base. The pKU20 contains structural gene for aminoglycoside phos-photransferase (aph), without promoter, and a multiple cloning site upstream the aph. Using this vector, a 412base pairs (bp) PstI fragment showing strong promoter activity both in Escherichia coli LE392 and Pseudomonas putida KCTC1644 has been cloned from Pseudomonas fluorescens chromosomal DNA on the basis of streptomycin resistance. The nucleotide sequence of the 412 bp fragment has been determined and the putative - 35 and -10 region was observed. Insecticidal protein gene of Bacillus thuringiensis subsp. kurstaki HD-73 inserted on downstream of the promoterlike DNA fragment was efficiently expressed in E. coli and P. putida. The toxin protein was efficiently synthesized in an insoluble form in both strains.

• Journal of Microbiology and Biotechnology
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• 제11권1호
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• pp.85-91
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• 2001

To construct an efficient Bacillus subtilis expression vector, strong promoters were isolated from the chromosomal DNA libraries of Clostridium acetobutylicum ATCC 4259, Thermoactinomyces sp. E79, and Bacillus thermoglucosidasius KCTC 3400. The $P_{C27}$ promoter cloned from the clostridial chromosmal DNA showed a 5-fold higher promoter strength than the $P_{SP02}$ promoter in the expression of the cat gene, and its sequence was estimated as an upstream region of the predicted hypothetical gene (tet-R family bacterial transcription regulator gene) in C. acetobutylicum. As a promoter element, $P_{C27}$ exhibited putative nucleotide sequences that can bind with bacterial RNAP and the 3'end of the 16S rRNA just upstream of the start codon. In addition, the promoter activity of $P_{C27}$ was distinctively repressed in the presence of glucose. Using $P_{C27}$ as the promoter element, a glucose controllable B. subtilis expression vector was constructed and the lipase gene from Staphylococcus haemolyticus KCTC 8957P was expressed in B. subtilis. When compared with the lipase expression by the T7 promoter induced by IPTG in E. coli, the $P_{C27}$ promoter showed about a 1.5-fold higher expression level in B. subtilis than that without induction.

• Journal of Microbiology
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• 제44권5호
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• pp.515-522
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• 2006

A cDNA library of Ganoderma lucidum has been constructed using a Zap Express cloning vector. A glyceraldehyde-3-phosphate dehydrogenase gene (gpd) was isolated from this library by hybridization of the recombinant phage clones with a gpd-specific gene probe generated by PCR. By comparison of the cDNA and the genomic DNA sequences, it was found that the complete nucleotide sequence encodes a putative polypeptide chain of 338 amino acids interrupted by 6 introns. The predicted amino acid sequence of this gene shows a high degree of sequence similarity to the GPD proteins from yeast and filamentous fungi. The promoter region contains a CT-rich stretch, two CAAT boxes, and a consensus TATA box. The possibility of using the gpd promoter in the construction of new transformation vectors is discussed.

• 미생물학회지
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• 제30권6호
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• pp.488-494
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• 1992

Three clones containing mouse CD4 gene were prepared using AKR genomic cosmid library. The role of 6, 500 bp 5' flanking region of the first exon of the AKR CD4 gene in tissue or developmental stage specific expression of CD4 has been studied. The deletion constructs containing various amounts of CD4 5' flanking sequences were prepared, and they were transfected into the cell lines representing different cell types or developmental stages of CD4 expression. Study of the reporter gene expression revealed that at least 1, 700 bp of 5' flanking region did retain promoter activity for CD4 expression. This area did not seem to contain enhancer activity for a full expression of CD4. However, the putative promoter interacted with other tissue specific enhancer sequence and showed the tissue specificity of the enhancer element.

• 한국축산식품학회지
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• 제31권3호
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• pp.381-388
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• 2011

Dietary conjugated linoleic acid (CLA) play key roles in lipid metabolism. Here, we investigated the effect of CLA on the transcriptional activity of TR4, an orphan nuclear receptor that plays an important role in lipid homeostasis. CLA increased TR4 gene mRNA level in 3T3-L1 adipocytes, but inhibited TR4 transcriptional activity in a dose-dependent manner. TR4 induced perilipin expression in 3T3-L1 adipocytes by activating perilipin promoter activity. In a gel shift assay, TR4 bound direct to the putative TR4 response element in the perilipin promoter. Interestingly, CLA reduced the interaction between TR4 and consensus DR1, a well-known TR4 binding site. Additionally, CLA inhibited TR4-induced perilipin promoter activity in a dose-dependent manner. Together, our results suggest that CLA may play a role in lipid homeostasis in adipocytes by functionally regulating TR4.