• Proceedings of the IEEK Conference
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• 2003.07d
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• pp.1569-1572
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• 2003

최근 생명공학 분야의 기술이 혁신적으로 발달함에 따라 게놈 프로젝트가 본래 계획보다 2년 앞당겨져 2003 년 4 월 인간 유전자의 완전한 서열을 밝히고 성공적으로 완료됨으로서 관련 연구자들은 인간의 유전자에 대한 대량의 서열 데이터를 얻게 되었다. 그래서 게놈 프로젝트의 다음 단계로서 엄청난 양의서열 정보 분석으로부터 유전자의 기능을 파악하고자 하는 연구들이 이미 세계적으로 활발히 진행되고 있다. 이러한 연구들의 최종적 목표는 질병 치료와 생명연장의 실현이라고 볼 수 있다. 유전자 연구를 위해선 우선 일차적으로 유전자 부위를 파악해야 한다. 유전자는 구조적으로 다시 여러 부분으로 나뉘는데 유전자 발현의 개시에 매우 중요한 요소 중 하나가 바로 프로모터 (Promoter) 이다. 프로모터 내에는 TATA box 가 있는데 이는 프로모터의 핵심 요소이다. 프로모터는 생명체의 종 그리고 RNA 중합효소의 종류에 따라 다르다. 이 논문에서는 다양한 신경망 알고리즘 중의 하나인 Backtpropagation 을 이용하여 밝혀지지 알은 서열에서 인간을 포함하는 원핵생물의 프로모터 서열을 예측할 수 있는 방법을 얻었기에 소개하고자 한다.

• Animal cells and systems
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• v.2 no.2
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• pp.287-295
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• 1998

The Saccharomyces cerevisiae MGMT gene encodes a O6-methylguanine DNA methyltransferase that protects cells from mutation or death by DNA alkylating agents. Using an in vitro transcription system, we analyzed its promoter region to find regulatory elements for transcription initiation. DNase I footprinting and a transcription assay showed that a functional TATA box, 5'-TGATATAGCA-3', is located in the region spanning from -25 to -34. We also found one upstream repressing sequence (URS), -333 to -213, by promoter deletion and competition analysis. Gel mobility shift assays and Southwestern blot analysis using URS region indicate specific complex formations. These results indicate that several cis-acting and trans-acting elements might be involved in the transcriptional regulation of the S. cerevisiae MGMT gene.

• Proceedings of the Korean Information Science Society Conference
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• 2003.10b
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• pp.769-771
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• 2003

PromSearch는 DNA 염기서열 상에서 프로모터의 위치를 예측하는 프로그램이다. 다루는 대상은 인간 DNA의 프로모터이며, 프로모터의 TSS(transcription start site, 전사시작지점)를 예측하는 것을 목표로 한다. 프로모터 영역을 세분하여 각 영역에 대한 프로파일을 PWM(position weight matrix)을 이용해 작성하며, 임의의 염기서열이 입력으로 주어지면 세분한 영역의 점수를 신경망을 이용해 통합하여 프로모터 여부와 TSS의 위치를 결정한다. 프로모터 영역의 분할은 코어 프로모터의 구성 요소인 TATA-box와 Inr, DPE(downstream promoter element), 그리고 코어 프로모터의 위쪽으로 150bp 크기의 영역 등으로 4분할하였다. Fickett의 데이터를 이용한 평가 결과 sensitivity 43%, specificity 88fp(1/376bp)의 성능을 보였다.

• Journal of Microbiology
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• v.33 no.3
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• pp.196-200
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• 1995

In order to express the CDC70 gene of Saccharomyces cerevisiae by heat shock, we have designed heat inducibe hybrid promoters using the Drosophila melanogaster heat shock elements (HSEs). A 220 bp-long upstream fragment of the D. melanogaster hsp70 gene comprised of four HSEs was placed upstream of the putative proximal TATA box of the CDC70 gene. Hybrid promoters containing different fusion joints were tested for their ability to drive the CDC70 gene expression by heat shock. The results showed that the HSEs of D. melanogaster conferred the heat-induced CDC70 gene expression, but the heat inducibility was much lower than that in D. melanogaster.

• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.19-19
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• 2001

Mammalian urothelium undergoes unique membrane specialization by making the asymmetric unit membrane (AUM) that is covered with the apical cell surface during terminal differentiation. The AUM contains several major integral membrane proteins including uroplakin Ia, Ib, II and III. The genes for uroplakins have been cloned from humans and mice, but not from porcine. In this study, we report the cloning of the UPII genomic DNA, which codes for the full length open reading frame for the uroplakin II protein. The deduced amino acid sequence encodes of a hydrophobic NH$_2$-terminal peptide, a prosequence, and a mature protein. The prosequence contains three potential N-glycosylation sites and a RGRR cleavage site that may be involved in uroplakin II processing and maturation. Northern and immunohistochemistry analyses showed that the porcine UPII gene is only expressed in urothelium and that the protein was specifically localized in urothelial superficial cells. A 2kb of upstream in the promoter sequence contains multiple transcription factor binding sites, including GC-box, SPI, AP2, and GATA-box sites, but not for TATA or CAAT-box sequences. Comparison of the porcine UPII promoter sequence with that of the murine by MEME system presented two conserved motifs, suggesting a cis-acting regulatory role for the conserved sequences. Sequence homology between two species in motif A and B was 79% and 80% respectively, although their relative locations were different. During the gestation, mouse bladder at estrus stages and day 10 after parturition showed higher UPII expression, while showed lower expression at peri-implantation stage. Taken together, our results showed that the porcine UPII gene was expressed highly and specifically in the bladder urothelium and that steroid hormones for implantation changed the expression of UPII in the bladder, although the biological significance of UPII remains to be not determined.

• Journal of Life Science
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• v.15 no.5 s.72
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• pp.802-808
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• 2005

Human Disabled-2 (Dab2) is a candidate tumor suppressor gone that regulates cell growth by c-Fos suppression in normal cells. In many cancer cells, Dab2 expression is lost or greatly diminished in $\∼85\%$ of the breast and ovarian cancers. In this study, we have examined the methylation status of CpG island on Dab2 gene promoter using bisulfite-assisted genomic sequencing and methylation specific PCR (MSP) method in human breast cancer cell line, MDA MB-231 cells. In normal human uterus endometrial cells, Dab2 was completely unmethylated. In contrast, Dab2 was methylated on CpG dinucleotides near the TATA_ box in MDA MB-231 cells. following MDA MB-231 cells by treatment with 5-azacytidine, Dab2 gene were demethylated and reexpressed. Result of this study suggested that silencing of Dab2 gene is correlated to CpG island methylation in human breast cancer cell line, MBA MD-231 cells.

• Asian-Australasian Journal of Animal Sciences
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• v.33 no.12
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• pp.2021-2030
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• 2020

Objective: Quantitative polymerase chain reaction (qPCR) has been extensively used in the field of mesenchymal stem cell (MSC) research to elucidate their characteristics and clinical potential by normalization of target genes against reference genes (RGs), which are believed to be stably expressed irrespective of various experimental conditions. However, the expression of RGs is also variable depending on the experimental conditions, which may lead to false or contradictory conclusions upon normalization. Due to the current lack of information for a clear list of stable RGs in bovine MSCs, we conducted this study to identify suitable RGs in bovine MSCs. Methods: The cycle threshold values of ten traditionally used RGs (18S ribosomal RNA [18S], beta-2-microglobulin [B2M], H2A histone family, member Z [H2A], peptidylprolyl isomerase A [PPIA], ribosomal protein 4 [RPL4], succinate dehydrogenase complex, subunit A [SDHA], beta actin [ACTB], glyceraldehyde-3-phosphate dehydrogenase [GAPDH], TATA box binding protein [TBP], and hypoxanthine phosphoribosyltrasnfrase1 [HPRT1]) in bovine bone marrow-derived MSCs (bBMMSCs) were validated for their stabilities using three types of RG evaluation algorithms (geNorm, Normfinder, and Bestkeeper). The effect of validated RGs was then verified by normalization of lineage-specific genes (fatty acid binding protein 4 [FABP4] and osteonectin [ON]) expressions during differentiations of bBMMSCs or POU class 5 homeobox 1 (OCT4) expression between bBMMSCs and dermal skins. Results: Based on the results obtained for the three most stable RGs from geNorm (TBP, RPL4, and H2A), Normfinder (TBP, RPL4, and SDHA), and Bestkeeper (TBP, RPL4, and SDHA), it was comprehensively determined that TBP and RPL4 were the most stable RGs in bBMMSCs. However, traditional RGs were suggested to be the least stable (18S) or moderately stable (GAPDH and ACTB) in bBMMSCs. Normalization of FABP4 or ON against TBP, RPL4, and 18S presented significant differences during differentiation of bBMMSCs. However, although significantly low expression of OCT4 was detected in dermal skins compared to that in bBMMSCs when TBP and RPL4 were used in normalization, normalization against 18S exhibited no significance. Conclusion: This study proposes that TBP and RPL4 were suitable as stable RGs for qPCR study in bovine MSCs.

• The Korean Journal of Physiology and Pharmacology
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• v.1 no.2
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• pp.195-200
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• 1997

Water transport in highly-permeable membranes is facilitated by some specialized pathways, which are called aquaporins (AQP). AQP1 (AQP-CHIP) is the first recognized aquaporin identified from red cells and renal proximal tubules. Up until now 4 other aquaporin homologs have been reported. Each aquaporin has its unique tissue distribution and regulatory mechanims. To elucidate molecular mechanisms for their transcription regulation and tissue-specific expression isolation of aquaporin genes is required. To clone promoters of the AQP family mouse genomic library was screened by the 1st exon-specific probe of AQP4, and 5 different plaques were positively hybridized. Phage DNAs were purified and characterized by restriction mapping and sequencing. One of them is the mouse AQP-CD gene. The gene was consisted of 4 exons and the exon-intron boundaries of mouse AQP-CD gene were identified at identical positions in other related genes. The 5'-flanking region of AQP-CD gene contains one classic TATA box, a GATA consensus sequence, an E-box and a cyclic AMP-responsive element. The cloning of the mouse AQP-CD gene, of which product is expressed in the collecting duct and is responsible for antidiuresis by vasopressin, will contribute to understand the molecular mechanisms of tissue-specific expression and regulation of AQP-CD gene under various conditions.

• Journal of Microbiology and Biotechnology
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• v.22 no.9
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• pp.1177-1184
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• 2012

In order to estimate how diverse the mating types in Pleurotus eryngii from different regions are, pairings between monokaryons derived from inter- and intra-groups were done. Sixteen and 15 alleles were identified at loci A and B from the 12 strains. In the P. eryngii KNR2312, widely used for commercial production, four mating loci, A3, A4, B3, and B4, were determined. Those loci, except A3, were found in 4 strains out of 12 strains. To improve breeding efficiency, especially in mating type determination, RAPD and BSA were performed to screen for a mating type specific marker. The SCAR marker 13-$2_{2100}$ was developed based on the RAPD-derived sequence typing B3 locus. The sequence analysis of 13-$2_{2100}$ revealed that it contained a conserved domain, the STE3 super-family, and consensus sequences like the TATA box and GC box. It seems likely that the SCAR marker region is a part of the pheromone receptor gene.

• BMB Reports
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• v.29 no.5
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• pp.468-473
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• 1996

The P38 promoter of minute virus of mice (MVM) is a very weak promoter which is strongly transactivated by viral nonstructural proteins. To analyze the upstream sequence of the P38 promoter which is responsible for the transactivation by nonstructural proteins in MVM, chloramphenicol acetyltransferase (CAT) reporter plasm ids containing a series of 5' deletion and internal deletion mutants of the P38 promoter were constructed. The wild type and mutant CAT constructs of P38 promoter were cotransfected into murine A92L fibroblast cells with a plasmid expressing viral nonstructural proteins by DEAE-dextran method. Each promoter activity was analyzed by CAT assay. As previously reported (Ahn et al., 1992), the proximal DNA cis-elements required for transactivation of the MVM P38 promoter are GC box and TATA box. However, the analysis of 5' deletion mutants showed that H-l tar like sequence (MVM TAR) which is located between -143 and -122 relative to the transcription initiation site is also required for transactivation of the P38 promoter by nonstructural proteins. Interestingly, even if the MVM TAR was removed by internal deletion, the level of the transactivation is still 70% of wild type level of transactivation. We also found that, in addition to the MVM TAR motif, there are two other motifs which are similar to the MVM TAR sequence. When these TAR like motifs were further deleted, the levels of transactivation were decreased further. Taken together, the MVM TAR sequence and TAR like motifs located upstream of P38 promoter are playing an important role for the transactivation of P38 promoter by nonstructural proteins in minute virus of mice.