• Molecules and Cells
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• 제41권11호
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• pp.953-963
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• 2018

The stepwise development of T cells from a multipotent precursor is guided by diverse mechanisms, including interactions among lineage-specific transcription factors (TFs) and epigenetic changes, such as DNA methylation and hydroxymethylation, which play crucial roles in mammalian development and lineage commitment. To elucidate the transcriptional networks and epigenetic mechanisms underlying T-cell lineage commitment, we investigated genome-wide changes in gene expression, DNA methylation and hydroxymethylation among populations representing five successive stages of T-cell development (DN3, DN4, DP, $CD4^+$, and $CD8^+$) by performing RNA-seq, MBD-seq and hMeDIP-seq, respectively. The most significant changes in the transcriptomes and epigenomes occurred during the DN4 to DP transition. During the DP stage, many genes involved in chromatin modification were up-regulated and exhibited dramatic changes in DNA hydroxymethylation. We also observed 436 alternative splicing events, and approximately 57% (252) of these events occurred during the DP stage. Many stage-specific, differentially methylated regions were observed near the stage-specific, differentially expressed genes. The dynamic changes in DNA methylation and hydroxymethylation were associated with the recruitment of stage-specific TFs. We elucidated interactive networks comprising TFs, chromatin modifiers, and DNA methylation and hope that this study provides a framework for the understanding of the molecular networks underlying T-cell lineage commitment.

• Molecular & Cellular Toxicology
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• 제3권3호
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• pp.190-194
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• 2007

Gastric adenocarcinoma (GA) is a major tumor type of gastric cancers and subdivides into several different tumors such as papillary, tubular mucinous, signet-ring cell and adenosquamous carcinoma according to histopatholigical determination. In other hand, GA is also subdivided into intestinal and diffuse type of adenocarcinoma by the Lauren?fs classification. In this study, we have examined differential gene expression pattern analysis of three histologically different GAs of 24 samples by using DNA microarray containing approximately 19000 genetic elements. The hierarchical clustering analysis of 24 gastric adenocarcinomas (12 of intestinal type, 7 of diffuse type and 5 of mixed type) resulted in two major subgroup on dendrogram, and two subgroups included most of intestinal and diffused type of GAs respectively. Supervised analysis of 19 intestinal and diffuse type GAs by using Wilcoxon rank T-test (P<0.01) resulted in 100 outlier genes which exactly separated intestinal and diffuse type of GA by differential gene expression. In conclusion, genome-wide analysis of gene expression of GAs suggested that GAs may subclassify as intestinal and diffused type of GA by their characteristic molecular expression. Our results also provide large-scale genetic elements which reflect molecular differences of intestinal and diffuse type of GAs, and this may facilitate to understand different molecular carcinogenesis of gastric cancer.

• Asian-Australasian Journal of Animal Sciences
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• 제31권1호
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• pp.138-148
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• 2018

Objective: Fusarium mycotoxins deoxynivalenol (DON) and zearalenone (ZEN), common contaminants in the feed of farm animals, cause immune function impairment and organ inflammation. Consequently, the main objective of this study was to elucidate DON and ZEN effects on the mRNA expression of pro-inflammatory cytokines and other immune related genes in the kidneys of piglets. Methods: Fifteen 6-week-old piglets were randomly assigned to three dietary treatments for 4 weeks: control diet, and diets contaminated with either 8 mg DON/kg feed or 0.8 mg ZEN/kg feed. Kidney samples were collected after treatment, and RNA-seq was used to investigate the effects on immune-related genes and gene networks. Results: A total of 186 differentially expressed genes (DEGs) were screened (120 upregulated and 66 downregulated). Gene ontology analysis revealed that the immune response, and cellular and metabolic processes were significantly controlled by these DEGs. The inflammatory stimulation might be an effect of the following enriched Kyoto encyclopedia of genes and genomes pathway analysis found related to immune and disease responses: cytokine-cytokine receptor interaction, chemokine signaling pathway, toll-like receptor signaling pathway, systemic lupus erythematosus (SLE), tuberculosis, Epstein-Barr virus infection, and chemical carcinogenesis. The effects of DON and ZEN on genome-wide expression were assessed, and it was found that the DEGs associated with inflammatory cytokines (interleukin 10 receptor, beta, chemokine [C-X-C motif] ligand 9, CXCL10, chemokine [C-C motif] ligand 4), proliferation (insulin like growth factor binding protein 4, IgG heavy chain, receptor-type tyrosine-protein phosphatase C, cytochrome P450 1A1, ATP-binding cassette sub-family 8), and other immune response networks (lysozyme, complement component 4 binding protein alpha, oligoadenylate synthetase 2, signaling lymphocytic activation molecule-9, ${\alpha}$-aminoadipic semialdehyde dehydrogenase, Ig lambda chain c region, pyruvate dehydrogenase kinase, isozyme 4, carboxylesterase 1), were suppressed by DON and ZEN. Conclusion: In summary, our results indicate that high concentrations of DON and ZEN suppress the inflammatory response in kidneys, leading to potential effects on immune homeostasis.

• Asian Pacific Journal of Cancer Prevention
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• 제17권9호
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• pp.4517-4525
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• 2016

Background: Sperm DNA damage is underlying aetiology of poor implantation and pregnancy rates but also affects health of offspring and may also result in denovo mutations in germ line and post fertilization. This may result in complex diseases, polygenic disorders and childhood cancers. Childhood cancer like retinoblastoma (RB) is more prevalent in developing countries and the incidence of RB has increased more than three fold in India in the last decade. Recent studies have documented increased incidence of cancers in children born to fathers who consume alcohol in excess and tobacco or who were conceived by assisted conception. The aetiology of childhood cancer and increased disease burden in these children is lin ked to oxidative stress (OS) and oxidative DNA damage( ODD) in sperm of their fathers. Though several antioxidants are in use to combat oxidative stress, the effect of majority of these formulations on DNA is not known. Yoga and meditation cause significant decline in OS and ODD and aid in regulating OS levels such that reactive oxygen speues meditated signal transduction, gene expression and several other physiological functions are not disrupted. Thus, this study aimed to analyze sperm ODD as a possible etiological factor in childhood cancer and role of simple life style interventions like yoga and meditation in significantly decreasing seminal oxidative stress and oxidative DNA damage and thereby decreasing incidence of childhood cancers. Materials and Methods: A total of 131 fathers of children with RB (non-familial sporadic heritable) and 50 controls (fathers of healthy children) were recruited at a tertiary center in India. Sperm parameters as per WHO 2010 guidelines and reactive oxygen species (ROS), DNA fragmentation index (DFI), 8-hydroxy-2'-deoxy guanosine (8-OHdG) and telomere length were estimated at day 0, and after 3 and 6 months of intervention. We also examined the compliance with yoga and meditation practice and smoking status at each follow-up. Results: The seminal mean ROS levels (p<0.05), sperm DFI (p<0.001), 8-OHdG (p<0.01) levels were significantly higher in fathers of children with RB, as compared to controls and the relative mean telomere length in the sperm was shorter. Levels of ROS were significantly reduced in tobacco users (p<0.05) as well as in alcoholics (p<0.05) after intervention. DFI reduced significantly (p<0.05) after 6 months of yoga and meditation practice in all groups. The levels of oxidative DNA damage marker 8-OHdG were reduced significantly after 3 months (p<0.05) and 6 months (p<0.05) of practice. Conclusions: Our results suggest that OS and ODD DNA may contribute to the development of childhood cancer. This may be due to accumulation of oxidized mutagenic base 8OHdG, and elevated MDA levels which results in MDA dimers which are also mutagenic, aberrant methylation pattern, altered gene expression which affect cell proliferation and survival through activation of transcription factors. Increased mt DNA mutations and aberrant repair of mt and nuclear DNA due to highly truncatred DNA repair mechanisms all contribute to sperm genome hypermutability and persistant oxidative DNA damage. Oxidative stress is also associated with genome wide hypomethylation, telomere shortening and mitochondrial dysfunction leading to genome hypermutability and instability. To the best of our knowledge, this is the first study to report decline in OS and ODD and improvement in sperm DNA integrity following adoption of meditation and yoga based life style modification.This may reduce disease burden in next generation and reduce incidence of childhood cancers.

• 응용통계연구
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• 제22권4호
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• pp.781-792
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• 2009

유전체 초기단계 연구에서는 비교적 소수의 마이크로어레이 샘플자료로서 실험을 진행하여 심도 깊게 연구해야 할 유전자 부분군(subsets)을 탐색하게 된다. 이러한 과정에서 요구되는 부분군 탐색에 사용되는 분석방법은 다수 샘플자료 분석의 경우와는 매우 다른 방법들이다. 유전자 극소수 샘플자료의 분석에 매우 적절한 방법인 랜덤검정법을 적용하여 정확한 P값(exact P value)의 이산형 분포가 얻어지고, 일양분포 귀무가설의 검정으로 유의 유전자가 존재하는지를 파악할 수 있다. 한 단계 더 나아가 Fuchs와 Kenett (1980)이 제시한 M 검정을 이용하여 이산형 P 값 다항분포에서 이상범주군(outlier cells)을 찾을 수 있으며 이로써 유의 유전자로서의 가능성이 있는 유전자군을 선정한다. 대다수의 마이크로어레이 유전체 연구에서 수 천 또는 수 만개의 유전자가 서로 독립이라고 가정하고 분석하는 것이 문제점이다. 그러나 본 논문에서는 유전자 연관성을 그대로 유지하는 순열에 기초한 랜덤검정법과 M 검정법으로서 유전자 연관성이 분석에 미치는 영향을 모의실험으로 알아보았으며, 그 영향이 결코 미약하지 않음을 확인할 수 있었다.

• 한국정보통신학회:학술대회논문집
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• 한국정보통신학회 2018년도 춘계학술대회
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• pp.588-591
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• 2018

Baculovirus는 원래 알팔파 루퍼 (looper)로부터 분리되었으며 154 개의 오픈 리딩 프레임 (ORF)을 가진 134-kbp 게놈을 포함하고 있다. 주요 캡시드 단백질 VP39는 약간의 단백질과 함께 p6.9 단백질로 DNA를 감싸는 뉴클레오 캡시드($21nm{\times}260nm$)로 형성된다. 그것들은 막대 모양의 캡시드 안에 이중 가닥의 고리 모양의 슈퍼 코일 DNA 분자이다. 야생형 baculovirus는 용균 및 폐색 된 생명주기를 모두 나타내며 바이러스 복제의 3 단계에 걸쳐 독립적으로 발달한다. 재조합 baculovirus는 광범위한 포유류 세포 유형에서 벡터를 전달하고 재조합 단백질을 발현 할 수 있다. 특히, 이들 baculovirus 벡터에 우세한 선별 마커를 포함시킴으로써 많은 세포에서 다양한 재조합 유전자를 발현시킬 수 있다. 본 연구의 배큘로 바이러스 벡터는 cytomegalovirus (CMV) 프로모터, uroplakin II promoter, polyhedron promoter, 수포 구내염 바이러스 G (VSVG), 녹색 형광 단백질 (EGFP), 단백질 전달 도메인 (PTD) 유전자 등으로 재구성되었다. 이러한 재구성 된 벡터를 다양한 세포 및 세포주에 감염시켰다. 우리는 다른 재조합 벡터와 비교하여 이러한 재조합 벡터의 전이 및 발현을 조사하는 수행하였다. 본 연구에서, 우리는 이 재조합 벡터의 형질 감염 및 발현이 어떤 대조군 벡터보다 더 높은 효능을 갖는다는 것을 알았다. 본 연구는 과학 기술부, 한국 정보 기술 진흥 기금 (MSIP)이 후원하는 한국 연구 재단 (NRF)을 통해 중견 연구원 프로그램 (NRF-2016R1A2B4016552)을 통해 지원되었다.

• Genes and Genomics
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• 제40권11호
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• pp.1181-1197
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• 2018

Tropical plant rubber tree (Hevea brasiliensis) is the sole source of commercial natural rubber and low-temperature stress is the most important limiting factor for its cultivation. To characterize the gene expression profiles of H. brasiliensis under the cold stress and discover the key cold stress-induced genes. Three cDNA libraries, CT (control), LT2 (cold treatment at $4^{\circ}C$ for 2 h) and LT24 (cold treatment at $4^{\circ}C$ for 24 h) were constructed for RNA sequencing (RNA-Seq) and gene expression profiling. Quantitative real time PCR (qRT-PCR) was conducted to validate the RNA-Seq and gene differentially expression results. A total of 1457 and 2328 differentially expressed genes (DEGs) in LT2 and LT24 compared with CT were respectively detected. Most significantly enriched KEGG pathways included flavonoid biosynthesis, phenylpropanoid biosynthesis, plant hormone signal transduction, cutin, suberine and wax biosynthesis, Pentose and glucuronate interconversions, phenylalanine metabolism and starch and sucrose metabolism. A total of 239 transcription factors (TFs) were differentially expressed following 2 h or/and 24 h of cold treatment. Cold-response transcription factor families included ARR-B, B3, BES1, bHLH, C2H, CO-like, Dof, ERF, FAR1, G2-like, GRAS, GRF, HD-ZIP, HSF, LBD, MIKC-MADS, M-type MADS, MYB, MYB-related, NAC, RAV, SRS, TALE, TCP, Trihelix, WOX, WRKY, YABBY and ZF-HD. The genome-wide transcriptional response of rubber tree to the cold treatments were determined and a large number of DEGs were characterized including 239 transcription factors, providing important clues for further elucidation of the mechanisms of cold stress responses in rubber tree.

• Animal Bioscience
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• 제34권11호
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• pp.1739-1748
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• 2021

Objective: In recent years, long noncoding RNAs (lncRNAs) have been identified in many species, and some of them have been shown to play important roles in muscle development and myogenesis. However, the differences in lncRNAs between Kazakh cattle and Xinjiang brown cattle remain undefined; therefore, we aimed to confirm whether lncRNAs are differentially expressed in the longissimus dorsi between these two types of cattle and whether differentially expressed lncRNAs regulate muscle differentiation. Methods: We used RNA-seq technology to identify lncRNAs in longissimus muscles from these cattle. The expression of lncRNAs were analyzed using StringTie (1.3.1) in terms of the fragments per kilobase of transcript per million mapped reads values of the encoding genes. The differential expression of the transcripts in the two samples were analyzed using the DESeq R software package. The resulting false discovery rate was controlled by the Benjamini and Hochberg's approach. KOBAS software was utilized to measure the expression of different genes in Kyoto encyclopedia of genes and genomes pathways. We randomly selected eight lncRNA genes and validated them by quantitative reverse transcription polymerase chain reaction (RT-qPCR). Results: We found that 182 lncRNA transcripts, including 102 upregulated and 80 downregulated transcripts, were differentially expressed between Kazakh cattle and Xinjiang brown cattle. The results of RT-qPCR were consistent with the sequencing results. Enrichment analysis and functional annotation of the target genes revealed that the differentially expressed lncRNAs were associated with the mitogen-activated protein kinase, Ras, and phosphatidylinositol 3-kinase (PI3k)/Akt signaling pathways. We also constructed a lncRNA/mRNA coexpression network for the PI3k/Akt signaling pathway. Conclusion: Our study provides insights into cattle muscle-associated lncRNAs and will contribute to a more thorough understanding of the molecular mechanism underlying muscle growth and development in cattle.

• Journal of Microbiology and Biotechnology
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• 제18권4호
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• pp.631-638
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• 2008

Tandem affinity purification (TAP) method combined with LC-MS/MS is the most accurate and reliable way to study the interaction of proteins or proteomics in a genome-wide scale. For the first time, we used a TAP-tag as a mutagenic tool to disrupt protein interactions at the specific site. Although lots of commonly used mutational tools exist to study functions of a gene, such as deletional mutations and site-directed mutagenesis, each method has its own demerit. To test the usefulness of a TAP-tag as a mutagenic tool, we applied a TAP-tag to RNA polymerase II, which is the key enzyme of gene expression and is controlled by hundreds of transcription factors even to transcribe a gene. Our experiment is based on the hypothesis that there will be interrupted interactions between Pol II and transcription factors owing to the TAP-tag attached at the C-terminus of each subunit of Pol II, and the abnormality caused by interrupted protein interactions can be observed by measuring a cell-cycle of each yeast strain. From ten different TAP-tagged strains, Rpb7- and Rpb12-TAP-tagged strains show severe defects in growth rate and morphology. Without a heterodimer of Rpb4/Rpb7, only the ten subunits Pol II can conduct transcription normally, and there is no previously known function of Rpb7. The observed defect of the Rpb7-TAP-tagged strain shows that Rpb7 forms a complex with other proteins or compounds and the interruption of the interaction can interfere with the normal cell cycle and morphology of the cell and nucleus. This is a novel attempt to use a TAP-tag as a proteomic tool to study protein interactions.

• 응용통계연구
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• 제25권2호
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• pp.269-277
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• 2012

마이크로어레이 분석은 특이 발현하는 개별적인 유전자보다 유전자 온톨로지(Gene Ontology)와 같이 기능적 분류나 생물학적 경로(pathway)와 관련된 유전자군을 찾아내는 것이 그 해석의 용이성 때문에 최근 더욱 많은 연구가 진행되고 있다. 약물 처리에 의한 생물학적 반응을 연구할 때, 한 유전자군에 속하는 유전자들 각각의 특이 발현 여부의 유의성을 나타내는 $p$-value들을 취합하여 그 유전자군의 유의성을 결정하는 통계 검증 방법을 본 논문에서 소개하였다. 본 논문에 제시된 유전자군 분석(Gene group analysis) 방법은 Fisher's exact test나 permutation test와 같은 기존의 대표적인 방법들보다 더 정확하고 적용범위가 넓음을 실재 생물학 실험 자료의 분석을 통해 보였다. 제시된 유전자군 분석 방법은 SAS 프로그램으로 구현되었고 저자의 홈페이지(http://cafe.daum.net/go.analysis)에서 내려 받아 사용할 수 있다.