• Title, Summary, Keyword: RNAi

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RNA Interference: a Promising Therapy for Gastric Cancer

  • Felipe, Aledson Vitor;Oliveira, Juliana de;Chang, Paula Yun Joo;Moraes, Andrea Aparecida de Fatima Souza;Silva, Tiago Donizetti da;Tucci-Viegas, Vanina Monique;Forones, Nora Manoukian
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.14
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    • pp.5509-5515
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    • 2014
  • Gastric cancer (GC) remains a virtually incurable disease when metastatic and requires early screening tools for detection of early tumor stages. Therefore, finding effective strategies for prevention or recurrence of GC has become a major overall initiative. RNA-interference (RNAi) is an innovative technique that can significantly regulate the expression of oncogenes involved in gastric carcinogenesis, thus constituting a promising epigenetic approach to GC therapy. This review presents recent advances concerning the promising biomolecular mechanism of RNAi for GC treatment.

Genetically Engineered Mouse Models for Drug Development and Preclinical Trials

  • Lee, Ho
    • Biomolecules & Therapeutics
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    • v.22 no.4
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    • pp.267-274
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    • 2014
  • Drug development and preclinical trials are challenging processes and more than 80% to 90% of drug candidates fail to gain approval from the United States Food and Drug Administration. Predictive and efficient tools are required to discover high quality targets and increase the probability of success in the process of new drug development. One such solution to the challenges faced in the development of new drugs and combination therapies is the use of low-cost and experimentally manageable in vivo animal models. Since the 1980's, scientists have been able to genetically modify the mouse genome by removing or replacing a specific gene, which has improved the identification and validation of target genes of interest. Now genetically engineered mouse models (GEMMs) are widely used and have proved to be a powerful tool in drug discovery processes. This review particularly covers recent fascinating technologies for drug discovery and preclinical trials, targeted transgenesis and RNAi mouse, including application and combination of inducible system. Improvements in technologies and the development of new GEMMs are expected to guide future applications of these models to drug discovery and preclinical trials.

Expression of temperature responsive genes in cell cultures derived from Bombyx mori

  • Kim, Eun-Young;Kang, Min-Uk;Park, Kwan-Ho;Choi, Kwang-Ho;Nho, Si-Kab
    • International Journal of Industrial Entomology
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    • v.31 no.2
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    • pp.95-102
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    • 2015
  • Insects are heterotherms that exhibit a close relationship between their ecology (especially temperature changes) and physiology. In the present study, selected genes associated with cell death and temperature were examined to determine gene expression in Bombyx mori in high and low temperature environments. We determined the amount of dsRNA, different concentrations of dsRNA, and different type of cells to set the conditions most efficient for RNAi. We then prepared dsRNA transcripts of the genes associated with cell death and temperature response. We analyzed cell damage via Trypan blue staining and found that cell viability was reduced after knockdown of these genes. The special transduced cell lines produced in the present study can be applied in various research fields. We also expect that these cell lines can be used as a research tool for the precise functional analysis of various genes.

Mobile transposon-like element, clone MTi7: Finding its role(s) by RNA interference (Mobile transposon-like element, clone MTi7:RNA interference를 이용한 역할 규명)

  • Park, Chang-Eun;Shin, Mi-Ra;Jeon, Eun-Hyun;Cho, Sung-Won;Lee, Sook-Hwan;Kim, Kyung-Jin;Kim, Nam-Hyung;Lee, Kyung-Ah
    • Clinical and Experimental Reproductive Medicine
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    • v.30 no.4
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    • pp.299-307
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    • 2003
  • Objectives: The present study was conducted to evaluate the mobile transposon-like element, clone MTi7 (MTi7) expression in the mouse ovary and to determine its role(s) in the mouse oocytes by RNA interference (RNAi). Methods: MTi7 mRNA expression was localized by in situ hybridization in day5 and adult ovaries. Double stranded RNA (dsRNA) was prepared for c-mos, a gene with known function as control, and the MTi7. Each dsRNA was microinjected into the germinal vesicle (GV) stage oocytes then oocyte maturation and intracellular changes were evaluated. Results: In situ hybridization analysis revealed that MTi7 mRNA localized to the oocyte cytoplasm from primordial to preovulatory follicles. After dsRNA injection, we found 43-54% GV arrest of microinjected GV oocytes with 68%-90% decrease in targeted c-mos or MTi7 mRNA. Conclusions: This is the first report of the oocyte-specific expression of the MTi7 mRNA. From results of RNAi for MTi7, we concluded that the MTi7 is involved in the germinal vesicle breakdown in GV oocytes, and MTi7 may be implicated with c-mos for its function. We report here that RNAi provides an outstanding approach to study the function of a gene with unknown functions.

Host-Induced gene silencing of fungal pathogenic genes confer resistance to fungal pathogen, Magnaporthe Oryzae in rice

  • Jin, Byung Jun;Chun, Hyun Jin;Kim, Min Chul
    • Proceedings of the Korean Society of Crop Science Conference
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    • pp.134-134
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    • 2017
  • Recently, host-induced gene silencing (HIGS) system has been successfully applied into development of resistant crops against insects, fungal and viral pathogens. To test HIGS-mediated resistance in rice against rice blast fungus, Magnaporthe oryzae, we first tested possibility of movement of small non-coding RNA from rice cells to rice blast fungus. The rice blast fungus expressing GFP transgene were inoculated to transgenic rice plants ectopically expressing dsRNAi construct targeting fungal GFP gene. Expression of dsRNAi construct for GFP gene in transgenic plants significantly suppressed GFP expression in infected fungal cells indicating that small RNAs generated in plant cells can move into infected fungal cells and efficiently suppress the expression of fungal GFP gene. Consistent with these results, expression of dsRNAi constructs against 3 fungal pathogenic genes of M. oryzae in transgenic rice specifically and efficiently suppressed not only the expression of fungal pathogenic genes, but also fungal infection. The conidia of M. oryzae applied on leaf sheath of transgenic rice expressing dsRNAs against 3 fungal pathogenic genes showed abnormal development of primary hyphae and malfunction of appressorium, which is consistent with the phenotypes of corresponding fungal knock-out mutants. Taken these results together, here, we suggest a novel strategy for development of antifungal crops by means of HIGS system.

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Characterization of Expression of UV-Inducible Gene (UV100 and UV150) in Caenorhabditis elegans (Caenorhabditis elegans에서 분리한 자외선 유도유전자 (UV100과 UV150)의 발현 및 특성에 관한 연구)

  • Shin, Sue-Hwa;Choi, Eun-Young;Choi, In-Soon
    • Journal of Life Science
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    • v.16 no.4
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    • pp.704-709
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    • 2006
  • The present study intends to characterize the DNA damage-inducible responses in Caenorhabditis elegans. To study UV-inducible responses in C. elegans, two UV-inducible cDNA clones were isolated from C. elegans by using subtration hybridization method. To investigate the expression of isolated genes, UV100 and UV150, the cellular levels of the transcript were determined by Northern blot analysis after UV-irradiation. The transcripts of isolated gene increased rapidly and reached maximum accumulation after UV-irradiation. Compared to the message levels of control, the levels of maximal increase were approximately 2 folds to UV-irradiation. These results implied that the effects of damaging agents are complex and different regulatory pathways exist for the induction of these genes. To study the function of UV100 and UV150 gene in response to UV irradiation, we carried out a RNAi experiment and investigated the UV sensivity. This result indicated that UV100 gene involved in stage-specific repair pathway or regulated by development.

Changes in gene expression associated with oocyte meiosis after $Obox4$ RNAi

  • Lee, Hyun-Seo;Kim, Eun-Young;Lee, Kyung-Ah
    • Clinical and Experimental Reproductive Medicine
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    • v.38 no.2
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    • pp.68-74
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    • 2011
  • Objective: Previously, we found that oocyte specific homeobox (Obox) 4 plays significant role in completion of meiosis specifically at meiosis I-meiosis II (MI-MII) transition. The purpose of this study was to determine the mechanism of action of $Obox4$ in oocyte maturation by evaluating downstream signal networking. Methods: The $Obox4$ dsRNA was prepared by $in$ $vitro$ transcription and microinjected into the cytoplasm of germinal vesicle oocytes followed by $in$ $vitro$ maturation in the presence or absence of 0.2 mM 3-isobutyl-1-metyl-xanthine. Total RNA was extracted from 200 oocytes of each group using a PicoPure RNA isolation kit then amplified two-rounds. The probe hybridization and data analysis were used by Affymetrix Gene-Chip$^{(R)}$ Mouse Genome 430 2.0 array and GenPlex 3.0 (ISTECH, Korea) software, respectively. Results: Total 424 genes were up (n=80) and down (n=344) regulated after $Obox4$ RNA interference (RNAi). Genes mainly related to metabolic pathways and mitogen-activated protein kinase (MAPK) signaling pathway was changed. Among the protein kinase C (PKC) isoforms, PKC-alpha, beta, gamma were down-regulated and especially the MAPK signaling pathway PKC-gamma was dramatically decreased by $Obox4$ RNAi. In the cell cycle pathway, we evaluated the expression of genes involved in regulation of chromosome separation, and found that these genes were down-regulated. It may cause the aberrant chromosome segregation during MI-MII transition. Conclusion: From the results of this study, it is concluded that $Obox4$ is important upstream regulator of the PKC and anaphase-promoting complex action for maintaining intact germinal vesicle.

Physiological Characterization of an AtPGR from Arabidopsis Involved in Pathogen Resistance (애기장대 AtPGR 단백질의 병 저항성에 관한 생리적 특성 분석)

  • Chung, Moon-Soo;Kim, Cheol-Soo
    • Journal of Life Science
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    • v.21 no.9
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    • pp.1295-1300
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    • 2011
  • The AtPGR gene is induced by pathogen infection, jasmonic acid and salicylic acid treatment and may therefore play a role in plant defense responses. Arabidopsis thaliana Plasma membrane Glucose-responsive Regulator (AtPGR) was previously isolated from Arabidopsis, which confers glucose insensitivity on plants. To study its biological functions directly, we have characterized both loss-of-function RNAi mutant and gain-of-function transgenic overexpression plants for AtPGR in Arabidopsis. The AtPGR-overexpressing plants displayed enhanced resistance to a virulent strain of the bacterial pathogen Pseudomonas syringae as measured by a significant decrease in both bacterial growth and symptom development as compared to those in wild-type and RNAi plants. The enhanced resistance in the gain-of-function transgenic plants was associated with increased induction of SA-regulated PDF1.2 and JA-regulated PR1 by the bacterial pathogen. Thus, pathogen-induced AtPGR plays a positive role in defense responses to P. syringae.

Improving Cellulase Production in Trichoderma koningii Through RNA Interference on ace1 Gene Expression

  • Wang, Shao-Wen;Xing, Miao;Liu, Gang;Yu, Shao-Wen;Wang, Juan;Tian, Sheng-Li
    • Journal of Microbiology and Biotechnology
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    • v.22 no.8
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    • pp.1133-1140
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    • 2012
  • Ribonucleic acid interference (RNAi) inhibits the expression of target genes in a sequence-specific manner, and shows potential for gene knockdown in filamentous fungi, in which the locus-specific gene knockout occurs in low frequency. In this study, the function of the repressor of cellulase expression I (ACEI) was verified in Trichoderma koningii (T. koningii) YC01 through RNAi, and ace1-silenced strains with improved cellulase productivity were obtained. An expression cassette that transcribed the interfering double-stranded RNA (dsRNA) of ace1 was constructed and transformed into T. koningii, and the transformants, in which the expression of ace1 was successfully silenced, were selected. As a result of the ace1 gene silencing, the expression levels of the main cellulase and xylanase genes were elevated, and the enhanced production of total proteins, cellulase, and xylanase was observed in the cultivation. In addition, the down-regulation of ace1 resulted in an increasing expression of xyr1, but no clear variation in the expression of cre1, which suggested that ACEI acted as a repressor of the xyr1 transcription, but was not involved in the regulation of the cre1 expression. The results of this work indicate that ace1 is a valid target gene for enhancing enzyme production in T. koningii, and RNAi is an appropriate tool for improving the properties of industrial fungi.

Functional Genomic Approaches Using the Nematode Caenorhabditis elegans as a Model System

  • Lee, Jun-Ho;Nam, Seung-Hee;Hwang, Soon-Baek;Hong, Min-Gi;Kwon, Jae-Young;Joeng, Kyu-Sang;Im, Seol-Hee;Shim, Ji-Won;Park, Moon-Cheol
    • BMB Reports
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    • v.37 no.1
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    • pp.107-113
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    • 2004
  • Since the completion of the genome project of the nematode C. elegans in 1998, functional genomic approaches have been applied to elucidate the gene and protein networks in this model organism. The recent completion of the whole genome of C. briggsae, a close sister species of C. elegans, now makes it possible to employ the comparative genomic approaches for identifying regulatory mechanisms that are conserved in these species and to make more precise annotation of the predicted genes. RNA interference (RNAi) screenings in C. elegans have been performed to screen the whole genome for the genes whose mutations give rise to specific phenotypes of interest. RNAi screens can also be used to identify genes that act genetically together with a gene of interest. Microarray experiments have been very useful in identifying genes that exhibit co-regulated expression profiles in given genetic or environmental conditions. Proteomic approaches also can be applied to the nematode, just as in other species whose genomes are known. With all these functional genomic tools, genetics will still remain an important tool for gene function studies in the post genome era. New breakthroughs in C. elegans biology, such as establishing a feasible gene knockout method, immortalized cell lines, or identifying viruses that can be used as vectors for introducing exogenous gene constructs into the worms, will augment the usage of this small organism for genome-wide biology.