OLFM4 may mediate tumor growth via NF-kB pathway

김균언,이시여,서유리 충남대학교 생물공학연구소 2014 생물공학연구지 Vol.20 No.-

Nucleotide Sequence of the Bovine Gene for Follicle-Stimulating Hormone β-Subunit

KIM, KYOON E.,DAVID F. GORDON,RICHARD A. MAURER 충남대학교부설 생명공학연구소 1991 생물공학연구지 Vol.1 No.-

The gene for the β-subunit of follicle-stimulating hormone (FSH-β) was isolated from a library of bovine DNA fragments cloned in bacteriophage λ and the complete nucleotide sequence of the gene was determined. The bovine FSH-β gene contains approximately 4000 nucleotides and consists of three exons separated by two intervening sequences. The transcription initiation site of the gene was mapped by nuclease protection experiments. Analysis of RNA species present in pituitary mRNA demonstrated the presence of a 4.0-kb RNA containing FSH-β sequences, which is the appropriate size for the primary transcript of the gene. Comparison of nucleotide sequence of the 5′-flanking sequence of the FSH-β gene to the 5′-flanking regions of other pituitary glycoprotein hormone genes reveals little sequence similarity.

Cloning of a Rat μ(mu)Opioid Receptor Gene

설은영,김균언 한국유전학회 1995 Genes & Genomics Vol.17 No.4

A μ (mu) opioid receptor (μORc) chromosomal gene was cloned from a genomic library of rat liver using μ opioid receptor cDNA as a probe. Restriction mapping and Southern blot analysis with a probe corresponding to the 5′ untranslated and the portion of N-terminal region of μORc cDNA have confirmed that the clone contains at least 3 Kb of 5′ flanking region of the gene. Sequencing analysis also revealed a translation initiation codon ATG in the 500 bp DNA fragment out of this region. Currently, a series of 5′ deletion mutants are being prepared to localize regulatory elements of the gene in this region. Northern blot analysis demonstrated that μORc gene is expressed in the hypothalamus-derived GT1-1 cells but not in pituitary-derived GH₃ cells. Furthermore, cAMP was shown to have minor regulatory effect on the expression of the gene in proportion to the concentration and treated time.

Olfactomedin 4 Suppresses Tumor Growth and Metastasis of Mouse Melanoma Cells through Downregulation of Integrin and MMP Genes

박기선,김균언,김기광,Zheng-Hao Piao,Mi Kyung Kim,Hyun Jean Lee,Yong Chan Kim,이기성,Jeung-Hoon Lee 한국분자세포생물학회 2012 Molecules and cells Vol.34 No.6

Olfactomedin 4 (OLFM4) is highly expressed in gastroin-testinal cancers and has an anti-apoptotic function. The roles of OLFM4 in tumor growth and metastasis and how it functions in these processes remain elusive. We investigated the function of OLFM4 in tumor growth and metastasis using B16F10 mouse melanoma cells as an experimental system. Our results showed that OLFM4 had no positive effect on cell viability or cell cycle progression in B16F10 cells. However, it significantly suppressed the tumorigenicity of B16F10 cells, i.e., intradermal primary tumor growth and lung metastasis. OLFM4 also suppressed the migration and invasion of B16F10 cells in vitro. For further insight into the mechanisms underlying OLFM4-mediated suppression of tumor progression, we examined the effect of OLFM4 on the expression of integrin and matrix metalloproteinase (MMP), both of which are involved in tumor progression. Overexpression of OLFM4 clearly reduced the expression levels of integrin 1, integrin 4, integrin 5, integrin 6, and MMP9. Moreover, forced expression of MMP9 attenuated the inhibitory activity of OLFM4 on migration and invasiveness. Our findings provide the experimental evidence that OLFM4 may func-tion as a tumor suppressor and an anti-metastatic gene during tumor progression.

Zebrafish 발생 과정에서 chk1 의 기능 분석

김동선,김균언,장미숙,이명철 한국유전학회 2000 Genes & Genomics Vol.22 No.3

As an initial approach to examine the function of cell cycle checkpoint kinase 1(chk1) in embryonic development, a 1.461 Kb long cDNA encoding a zebrafish chk1(Zchk1) was isolated from the 12-day-old zebrafish cDNA library. DNA sequences of the cDNA shares 87% homology with human chk7 (hchk1). However, the cDNA contains an incomplete open reading frame lacking approximately 100 by at the 3'-end region. RT-PCR analysis showed that the level of Zchk1 mRNAs was present from 1 cell stage throughout the embryonic development. Whole mount in situ hybridization demonstrated that the mRNA persisted at low level through 50% epiboly, and became restricted in the anterior part of antero-posterior axis in 3 somites, and only in the tail region of primodia 11. Inhibition of the Zchk1 expression by microinjection of Zchk1 anti-sense RNA into 1-4 cell stage embryos caused retardation of embryonic development at late somite stage, which in turn resulted in dorsally rolled tail. Taken together, these results indicate that Zchk1 may be involved in the regulation of cell cycle during embryogenesis as well as in the formation of the dorsoventral axis in the posterior region.

Enhanced and sustained gene expression of Epstein - Barr virus - based plasmid with polyamidoamine dendrimer derivatives

최혜,박기선,김균언,최준식 한국고분자학회 2012 한국고분자학회 학술대회 연구논문 초록집 Vol.2012 No.4

Decreased Catalytic Subunit mRNA Levels and Altered Catalytic Subunit mRNA Structure in a cAMP-resistant Chinese Hamster Ovary Cell line

Howard, Paul,Day, Kathleen H.,Kim, Kyoon E.,Richardson, Jeanne,Thomas, James,Abraham,Irene,Fleischmann, Robert D.,Gottesman, Michael M.,Maurer, Richard A. 충남대학교부설 생명공학연구소 1992 생물공학연구지 Vol.2 No.-

The mechansims responsible for decreased levels of cAMP-dependent protein kinase activity in a mutant Chinese hamster ovary cell line have been examined. The cAMP-resistant Chinese hamster ovary 10260 cell line was found to possess only 20% of the cAMP-dependent protein kinase activity found in wild-type cells. The pressence of decreased concentrations of the catalytic subunit in these cells was confirmed through binding studies using a radiolabeled, heat-stable inhibitor of the kinase. Cloned Chinese hamster ovary catalytic subunit cDNAs were isolated, characterized, and used as hybridization probes to examine the relative concentrations of catalytic subunit mRNAs in the wild-type and 10260 cell lines. A 40-50% decrease in the concentration of the mRNA for the Cα isozyme of the catalytic subunit was observed in 10260 cells, as compared with wild-type. This decrease in catalytic subunit mRNA concentration probably accounts for a portion of the decreased kinase activity in the mutant cell. Further analysis of Cα mRNA by polymerase chain reaction confirmed the decreased expression of Cα mRNA in 10260 cells and further demonstrated the presence of two different species of Cα cDNAs was indistinguishable from the wild-type cDNA, but the other species was shorter. Nucleotide sequence analysis of the amplified cDNAs led to the identification of a 191-base pair deletion in the shorter cDNA. Gene transfer studies using wild-type and 10260 Cα cDNAs demonstrated wild-type activity, but the shorter cDNA was inactive. These studies suggest that at least two alternations in gene expression are responsible for decreased cAMP-dependent protein kinase activity in the 10260 cell line. One alteration results in an approximately 2-fold decrease in the concentrations of Cα mRNA in the cells.

사람의 성선자극 호르몬 α-Subunit 유전자와 결합하는 Trans - Acting Factor 의 확인 및 분석

백상기,송석빈,김균언 한국유전학회 1998 Genes & Genomics Vol.20 No.1

The 5' flanking region of the α-subunit gene of human chorionic gonadotropin was tested for the binding with the nuclear extracts of the JAR choriocarcinoma cells. Gel mobility shift assay has demonstrated that two BstNI restriction fragments, 150 bp and 340 bp respectively, have binding activities with the nuclear extracts. The 150 bp fragment was chosen to examine in detail, since several transcription factor binding sites have already been reported within the 340 bp fragment. Band competition assay showed that the binding of 150 bp fragment is highly sequence-specific. The binding activity was further narrowed down to a 55 bp fragment, which was generated by Nsil digestion of the 150 bp fragment. DNasel footprinting assay revealed the binding sites on the nucleotide levels from -511 to -499. The binding sites were again confirmed by a oligonucleotide-directed mutagenesis of the 150 bp fragment followed by a gel mobility shift assay. Functional aspect of this binding was evaluated with the DNA-mediated gene transfer techniques. However, in several approaches tried so far, no effect on the transcription efficiency was observed, implicating a structural role of the binding. In this line, it is worth to note that the binding activity of the 150 bp fragment was observed in every cell or tissue nuclear extracts tested so far.

흰쥐 μ(mu)opioid Receptor 유전자의 클로닝 및 발현 조절

설은영,송석빈,김균언 한국유전학회 1998 Genes & Genomics Vol.20 No.3

Chromosomal gene encoding a μ(mu) opioid receptor (μORc) was cloned from a rat genomic library using μORc cDNA as a probe. Sequence analysis of the clone identified a transcription start site, four AP-1 sites, two Oct-1 sites and one CREBP1 site in the immediate 5' flanking region as well as a translation initiation codon ATG in exon I. However, canonical TATA box was not found in the rat μORc promoter region. Northern blot analysis demonstrated that μORc gene is expressed in the hypothalamus-derived GT1-1 cells and pituitary gonadotrope-derived αT3-1 cells but neither in pituitary lactotrope-derived GH₃ cells nor in pituitary thyrotrope-derived αTSH cells. In addition, expression of the gene was induced in the cells treated with cAMP for 24 hrs. A series of deletion mutants of the 5'-flanking region were subcloned into the luciferase reporter vector, and tested for their transcriptional activities by transfecting into αT3-1 cells. The construct containing up to -2967 bp region exhibited maximum transcriptional activity among the tested constructs, whereas the region below -1705 bp conferred significantly low levels of activity. Therefore, it is conceivable that the region between -2967 bp and -1705 bp is responsible for the basal expression of the μORc gene.

LIM domain과 상호작용 하는 transcription cofactor hTAiL-1의 기능 분석

박경숙,송석빈,김균언 충남대학교 자연과학연구소 1999 忠南科學硏究誌 Vol.26 No.1

The LIM homeodomain (LIM-HD) proteins, which contain two tandem LIM domains followed by a homeodomain, are critical transcription factors for embryonic development. The LIM domain is a conserved cysteine-rich zinc-binding motif and mediate protein-protein interactions. We have isolated two transcription activator interacting with LIM domain (hTAiL-1 and -2) from human brain cDNA library on the basis of its ability to interact with the LIM-HD protein Lh-2 (or Lhx-2). Here we report on the function of the hTAiL-1 protein. Northern blot analysis using human tissue blot revealed that hTAiL-1 is very abundantly expressed in the heart and muscle. In order to confirm that hTAL-1 interacts with LIM domain, β-galactosidase assay using yeast two-hybrid system was firstly employed, followed by an in vitro protein interaction assay and a mammalian two-hybrid assay. The results of these assay demonstrated that hTAiL-1 interact with LIM domain in a very weak mode. In vivo functional data using transfection experiments in SL-2 cells suggested that Sp1 transcription factor may be involved in the interaction of hTAiL-1 with Lh-2. In fact, hTAiL-1 stimulates the promoter activity deriven by Sp1 more efficiently than by Lh-2. In addition, effect of hTAiL-1 can be observed in a series of transfection experiment with differentiated L6 cells. Taken together, these results indicated that hTAiL-1 may mediate specific protein-protein interactions between LIM-HD and additional Sp1 basal transcription factor.