• Journal of Life Science
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• v.26 no.1
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• pp.140-145
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• 2016

Genes in multicellular organisms are transcribed in development, differentiation, or tissue-specific manners. The transcription of genes is activated by enhancers, which are transcription regulatory elements located at long distances from the genes. Recent studies have reported that noncoding RNAs are transcribed from active enhancers by RNA polymerase II (RNA Pol II); these are called enhancer RNAs (eRNAs). eRNAs are transcribed bi-directionally from the enhancer core, and are capped on the 5’ end but not spliced or polyadenylated on the 3’ end. The transcription of eRNAs requires the binding of transcription activators on the enhancer and associates positively with the transcription of the target gene. The transcriptional inhibition of eRNAs or the removal of eRNA transcripts results in the transcriptional repression of the coding gene. The transcriptional procedure of eRNAs causes enhancer- specific histone modifications, such as histone H3K4me1/2. eRNA transcripts directly interact with Mediator and Rad21, a cohesin subunit, generating a chromatin loop structure between the enhancer and the promoter of the target gene. The recruitment of RNA Pol II into the promoter and its elongation through the coding region are facilitated by eRNAs. Here, we will review the features of eRNAs, and discuss the mechanism of eRNA transcription and the roles of eRNAs in the transcriptional activation of target genes.

• Korean journal of applied entomology
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• v.35 no.4
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• pp.315-320
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• 1996

The effects of 2-arnino-3-methyIimidazo[4,5-fq]u inoline (IQ), 2-amino-6dimethyl-dipyrido[l,2-a;3',2'-d] imidazole (Glu-P-1) and aflatoxin B1 (AFBI) on the mitotic recombinations and somatic chromosome mutations were investigated using the transgenic Drosophila bearing a chimeric gene consisting of a promoter region of Drosophila actin 5C gene and rat DNA polymerase $. For investigating mitotic recombinations and the somatic chromosome mutations, the heterozygous (mwhl+) strain possessing or lacking transgene pol P was used. The spontaneous frequency of small mwh spots, due to deletion or nondisjunction etc., in the non-transgenic w strain and the transgenic plpol $1-130 strain was 0.351 and 0.606, respectively. The spontaneous frequency (0.063) of large mwh spots, arising mostly from somatic recombination between the centromere and the locus mwh, in the transgenic plpol $1-130 strain, was about three times higher than that (0.021) of the non-transgenic w strain. The mutant clone frequencies of two types induced by two heterocyclic mines (IQ and Glu-P-1) and AFBl in the transformant pbol PI-130 were two or three times higher than those in the host strain w. These mean that rat DNA polymerase P participates at least in the somatic chromosome mutations and mitotic recombination processes. And the present results suggest that the transgenic Drosophl!~ used in this study can be used as a hypersensitive, in vivo short-term assaying system for various environmental mutagens.

• Korean Chemical Engineering Research
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• v.51 no.4
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• pp.500-505
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• 2013

In this study, we investigated the kinetics of gas hydrate formation in the presence of ionic liquid (IL). Hydroxyethyl-methyl-morpholinium chloride (HEMM-Cl) was chosen as a material for the promotion effect test. Phase equilibrium curve for $CH_4$ hydrate with aqueous IL solution was obtained and its induction time and consumed amount of $CH_4$ gas were also measured. Aqueous solutions containing 20~20,000 ppm of HEMM-Cl was prepared and studied at 70 bar and 274.15 K. To compare the measured results to those of the conventional promoter, sodium dodecyl sulfate was also tested at the same condition. Result showed that the hydrate equilibrium curve was shifted toward higher pressure and lower temperature region. In addition, the induction time on $CH_4$ hydrate formation in the presence of IL was not shown. The amount of consumed $CH_4$ was increased with the whole range of tested concentration of IL and the highest consumption of $CH_4$ happened at 1,000 ppm of HEMM-Cl. HEMM-Cl induced and enhanced the $CH_4$ hydrate formation with a small amount of addition. Obtained result is expected to be applied for the development of technologies such as gas storage and transport using gas hydrates.

• Journal of Life Science
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• v.22 no.12
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• pp.1600-1607
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• 2012

The matrix metalloproteinase (MMP) family plays essential roles in physiological processes such as embryonic development, angiogenesis, wound healing, and tissue homeostasis as a consequence of MMPr capacity for breaking down many types of extracellular matrix proteins. Imbalanced regulation of MMP expression can also lead to pathological conditions such as tumor progression. We recently reported that the Drosophila Mmp1 gene is highly expressed in the digestive tract and is required for the maintenance of intestinal homeostasis such as by restriction of uncontrolled intestinal stem cell proliferation. However, the regulatory mechanisms of MMP gene expression in the intestine remain unclear. In this study, we determined that the expression of Mmp1 is regulated by the homeodomain transcription factor Caudal. Experiments using the targeted expression of Caudal under the regulation of Gal4-UAS system indicated that endogenous Caudal is required for the Mmp1 gene expression in the adult Drosophila intestine and that exogenous Caudal induces Mmp1 expression. Transient transfection experiments indicated that Caudal can activate the promoter activity of Mmp1 and that several putative Caudal binding sites in the 5'-flanking region of the Mmp1 gene may be critical to the upregulation by Caudal. Our data suggest that Mmp1 is one of the target genes of Caudal in physiological normal condition and in tumorigenesis.

• Korean Journal of Animal Reproduction
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• v.26 no.2
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• pp.87-94
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• 2002

This study was performed during the four seasons for the production of transgenic pigs containing the Cellulase Digest Gene. Purebred Landrace gilts and sows approximately 8∼15 months of age (n=126) were used for the collection of 1-cell zygotes for DNA microinjection and transfer. Retrospectively, estrus synchronization and superovulation schemes were evaluated to assess practicality fur zygote collection. Synchronization and superovulation procedures were used that cyclic gilts were synchronized with 20mg altrenogest (ALT) per day for 9 days after PG600 administration followed by superovulation with 1000 IU pregnant mares serum gonadotropin (PMSG) and 750IU human chorionic gonadotrophin (hCG). The cellulase digestion gene for microinjection is rat elasterase promoter (rEl) linked to CelD gene. After hormone treatment, 1,422 embryos were collected from 91 donors and 95.6% (1,359/1,422) embryos were in 1-cell stage which can be visualized the pronuclei for DNA microinjection. A total of 725 DNA microinjected embryos transferred into 35 recipients and produced 65 piglets from 13 litters. Pregnancy rate according to the number of transferred embryos to recipients was higher the group which received 21 to 24 embryos (50.0%) than other groups 20.0% in less and 33.3% in more. A tail tissue was collected from 65 piglets for biopsy. PCR screening was performed on each DNA sample using two separate sets of primers specific for the 5'- and 3'-flanking region of the rEl-CelD gene. Five of the 65 piglets (7.69%) were positive for the transgene. This study provide useful information regarding production of transgenic pig for bioreactor research.

• Korean Journal of Microbiology
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• v.32 no.2
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• pp.102-108
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• 1994

Autonomously replicating sequence Binding Factor 1(ABF1) is a DNA-binding protein that specifically recognizes the $RTCRYN_5ACG$ at many sites in the yeast genome including the promoter element, mating-type silencer and ARS. To express the intact full-length ABF1 gene in E. coli, the ABF1 gene has been cloned into pMAL-c2 and His-61, Leu-353 and Leu-360 were substituted with other amino acid. ABF1 fusion proteins of wild type ABF1 and H61A, L353R and L360R nutants were purified by amylose resin affinity chromatography. Fusion protein of MBP and ABF1 was digested by Factor Xa and Characterized by gel retardation assay and complementation test. As aresult, we suggested that other DNA binding motif except atypical inc-finger motif is in the middle region of ABF1.

• Journal of agriculture & life science
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• v.45 no.6
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• pp.201-212
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• 2011

The alkaline protease gene was cloned from a halo-tolerant alkalophilic Bacillus clausii I-52 isolated from the heavily polluted tidal mud flat of West Sea in Inchon Korea, which produced a strong extracellular alkaline protease (BCAP). Based on the full genome sequence of Bacillus subtilis, PCR primers were designed to allow for the amplification and cloning of the intact pro-BCAP gene including promoter region. The full-length gene consists of 1,143 bp and encodes 381 amino acids, which includes 29 residues of a putative signal peptide and an additional 77-amino-acid propeptide at its N-terminus. The mature BCAP deduced from the nucleotide sequence consists of 275 amino acids with a N-terminal amino acid of Ala, and a relative molecular weight and pI value was 27698.7 Da and 6.3, respectively. The amino acid sequence shares the highest similarity (99%) to the nattokinase precursor from B. subtilis and subtilisin E precursor from B. subtilis BSn5. The substrate specificity indicated that the recombinant BCAP could hydrolyze efficiently the synthetic substrate, N-Suc-Ala-Ala-Pro-Phe-pNA,and did not hydrolyze the substrates with basic amino acids at the P1 site. The recombinant BCAP was strongly inhibited by typical serine protease inhibitor, PMSF, indicating that BCAP is a member of the serine proteases.

• BMB Reports
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• v.56 no.10
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• pp.569-574
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• 2023

Aberrant DNA methylation plays a pivotal role in the onset and progression of colorectal cancer (CRC), a disease with high incidence and mortality rates in Korea. Several CRC-associated diagnostic and prognostic methylation markers have been identified; however, due to a lack of comprehensive clinical and methylome data, these markers have not been validated in the Korean population. Therefore, in this study, we aimed to obtain the CRC methylation profile using 172 tumors and 128 adjacent normal colon tissues of Korean patients with CRC. Based on the comparative methylome analysis, we found that hypermethylated positions in the tumor were predominantly concentrated in CpG islands and promoter regions, whereas hypomethylated positions were largely found in the open-sea region, notably distant from the CpG islands. In addition, we stratified patients by applying the CpG island methylator phenotype (CIMP) to the tumor methylome data. This stratification validated previous clinicopathological implications, as tumors with high CIMP signatures were significantly correlated with the proximal colon, higher prevalence of microsatellite instability status, and MLH1 promoter methylation. In conclusion, our extensive methylome analysis and the accompanying dataset offers valuable insights into the utilization of CRC-associated methylation markers in Korean patients, potentially improving CRC diagnosis and prognosis. Furthermore, this study serves as a solid foundation for further investigations into personalized and ethnicity-specific CRC treatments.

• Journal of Microbiology
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• v.45 no.2
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• pp.158-167
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• 2007

In this study, we have cloned a novel cDNA encoding for a papain-family cysteine protease from the Uni-ZAP XR cDNA library of the polychaete, Periserrula leucophryna. This gene was expressed in Escherichia coli using the T7 promoter system, and the protease was characterized after partial purification. First, the partial DNA fragment (498 bp) was amplified from the total RNA via RT-PCR using degenerated primers derived from the conserved region of cysteine protease. The full-length cDNA of cysteine protease (PLCP) was prepared via the screening of the Uni-ZAP XR cDNA library using the $^{32}P-labeled$ partial DNA fragment. As a result, the PLCP gene was determined to consist of a 2591 bp nucleotide sequence (CDS: 173-1024 bp) which encodes for a 283-amino acid polypeptide, which is itself composed of an 59-residue signal sequence, a 6-residue propeptide, a 218-residue mature protein, and a long 3'-noncoding region encompassing 1564 bp. The predicted molecular weights of the preproprotein and the mature protein were calculated as 31.8 kDa and 25 kDa, respectively. The results of sequence analysis and alignment revealed a significant degree of sequence similarity with other eukaryotic cysteine proteases, including the conserved catalytic triad of the $Cys^{90},\;His^{226},\;and\;Asn^{250}$ residues which characterize the C1 family of papain-like cysteine protease. The nucleotide and amino acid sequences of the novel gene were deposited into the GenBank database under the accession numbers, AY390282 and AAR27011, respectively. The results of Northern blot analysis revealed the 2.5 kb size of the transcript and ubiquitous expression throughout the entirety of the body, head, gut, and skin, which suggested that the PLCP may be grouped within the cathepsin F-like proteases. The region encoding for the mature form of the protease was then subcloned into the pT7-7 expression vector following PCR amplification using the designed primers, including the initiation and termination codons. The recombinant cysteine proteases were generated in a range of 6.3 % to 12.5 % of the total cell proteins in the E. coli BL21(DE3) strain for 8 transformants. The results of SDS-PAGE and Western blot analysis indicated that a cysteine protease of approximately 25 kDa (mature form) was generated. The optimal pH and temperature of the enzyme were determined to be approximately 9.5 and $35^{\circ}C$, respectively, thereby indicating that the cysteine protease is a member of the alkaline protease group. The evaluation of substrate specificity indicated that the purified protease was more active towards Arg-X or Lys-X and did not efficiently cleave the substrates with non-polar amino acids at the P1 site. The PLCP evidenced fibrinolytic activity on the plasminogen-free fibrin plate test.

• Tuberculosis and Respiratory Diseases
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• v.51 no.2
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• pp.108-121
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• 2001

Background : The $p16^{INK4a}$ (p16) twnor suppressor gene is frequently inactivated in hwnan non-small cell lung cancers (NSCLCs), predominantly through homozygous deletion or in association with aberrant promotor hypermethylation. Death-associated protein kinase (DAPK) gene influences interferon $\gamma$-induced apoptotic cell death and has important role in metastasis of lung cancer in animal model. Hypermethylation of promoter region of DAP kinase gene may suppress the expression of this gene. Methods : This study was performed to investigate the aberrant methylation of p16 or DAP kinase in 35 resected primary NSCLCs by methylation-specific PCR (MSP), and demonstrated frequency, diagnostic value and clinical implication of aberrant methylation of two genes. Results : Thirty-two cases were male patients, and 3 cases were female patients with an average age was 57. $8{\pm}10.5$ years. The histologic types of lung cancer were 22 of squamous cell carcinoma, 12 of adenocarcinoma, 1 of large cell carcinoma. Pathologic stages were 11 cases of stage I (1 IA, 10 IB), 13 cases of stage II (1 IIA, 12 IIB), and 11 cases of stage III (9 IIIA, 2 IIIB). Regarding for the cancer tissue, p16 aberrant methylation was noted in 13 case of 33 cases (39.4%), DAP kinase in 21 cases of 35 cases (60%). Age over 55 year was associated with p16 aberrant methylation significantly (p<0.05). Methylation status of two genes was not different by smoking history, histologic type, size of tumor, lymph node metastasis and disease progression of lung cancer. There was no correlation between p16 and DAP kinase hypermethylation. Conclusion: This investigation demonstrates that aberrant methylation of p16 tumor suppressor gene or DAP kinase showed relatively high frequency (74.3%) in NSCLCs, and that these genes could be a biologic marker for early detection of lung cancer.