• Title/Summary/Keyword: Transcript System

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Identification of Oocyte-Specific Diva-Associated Proteins using Mass Spectrometry (Mass Spectrometry를 이용한 난자 특이적인 Diva와 상호작용하는 단백질의 동정)

  • Yoon, Se-Jin;Kim, Jung-Woong;Choi, Kyung-Hee;Lee, Sook-Hwan;Lee, Kyung-Ah
    • Clinical and Experimental Reproductive Medicine
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    • v.33 no.3
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    • pp.189-198
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    • 2006
  • Objective: We previously described that Diva is highly expressed in matured metaphase II (MII) oocytes compared to immature germinal vesicle (GV) oocytes in mouse. We report here that the expression of Diva transcript as well as protein is oocyte-specific. To elucidate its physiological role in oocyte, the binding partner(s) of Diva has been identified by using immunoprecipitation (IP) followed by Mass Spectrometry. Methods: NIH/3T3 cells were transiently transfected for 24 h with either empty vector for control or FLAG-tagged mouse Diva construct, and IP was performed with anti-FLAG antibody. The immuno-isolated complexes were resolved by SDS-PAGE on a 12% gel followed by Coomassie Blue staining. For in-gel digestion, 15 bands of interest were excised manually and digested with trypsin. All mass spectra were acquired at a positive reflector mode by a 4700 Proteomics Analyzer (Applied Biosystems, Framingham, MA). Proteins were identified by searching the NCBI nonredundant database using MASCOT Peptide Mass Fingerprint software (Matrixscience, London). Results: Diva-associated complexes were formed in FLAG-tagged mouse Diva-overexpressed NIH/3T3 cells via IP using anti-FLAG-conjugated beads. Among the excised 15 bands, actin and actin-binding proteins such as tropomyosin, tropomodulin 3, and ${\alpha}$-actinin were identified. Binding between Diva and actin or tropomyosin was confirmed by IP followed by Western blot analysis. Both bindings were also detected endogenously in mouse ovaries, indicating that Diva works with actin and tropomyosin. Conclusions: This is the first report that immuno-isolated Diva-associated complexes are related to actin filament of the cytoskeletal system. When we consider the association of Diva with actin and tropomyosin, oocyte-specific Diva may play a role in modulating the cytoskeletal system during oocyte maturation.

Cloning of a Glutathione S-Transferase Decreasing During Differentiation of HL60 Cell Line (HL6O 세포주의 분화 시 감소 특성을 보이는 Glutathione S-Transferase의 클로닝)

  • Kim Jae Chul;Park In Kyu;Lee Kyu Bo;Sohn Sang Kyun;Kim Moo Kyu;Kim Jung Chul
    • Radiation Oncology Journal
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    • v.17 no.2
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    • pp.151-157
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    • 1999
  • Purpose : By sequencing the Erpressed Sequence Tags of human 걸ermal papilla CDNA library, we identified a clone named K872 of which the expression decreased during differentiation of HL6O cell line. Materials and Methods : K872 plasmid DNA was isolated according to QIA plasmid extraction kit (Qiagen GmbH, Germany). The nucleotide sequencing was performed by Sanger's method with K872 plasmid DNA. The most updated GenBank EMBL necleic acid banks were searched through the internet by using BLAST (Basic Local Alignment Search Tools) program. Nothern bots were performed using RNA isolated from various human tissues and cancer cell lines. The gene expression of the fusion protein was achieved by His-Patch Thiofusicn expression system and the protein product was identified on SDS-PAGE. Results : K872 clone is 1006 nucleotides long, and has a coding region of 675 nucleotides and a 3' non-coding region of 280 nucleotides. The presumed open reading frame starting at the 5' terminus of K872 encodes 226 amino acids, including the initiation methionine residue. The amino acid sequence deduced from the open reading frame of K872 shares $70\%$, identity with that of rat glutathione 5-transferase kappa 1 (rGSTKl). The transcripts were expressed in a variety of human tissues and cancer cells. The levels of transcript were relatively high in those tissues such as heart, skeletal muscle, and peripheral blood leukocyte. It is noteworthy that K872 was found to be abundantly expressed in coloreetal cancer and melanoma cell lines. Conclusion : Homology search result suggests that K872 clone is the human homolog of the rGSTK1 which is known to be involved in the resistance of cytotoxic therapy. We propose that meticulous functional analysis should be followed to confirm that.

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Differential Effects of Acute and Chronic Exercise on Autophagy-related Gene Expression in Drosophila melanogaster (일회성 및 만성적 유산소운동이 초파리의 자가포식 관련 유전자 발현에 미치는 영향)

  • Kim, Hee Yeon;Kim, Hye Jin;Hwang, Ji Sun;Lee, Won Jun
    • Journal of Life Science
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    • v.24 no.11
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    • pp.1180-1186
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    • 2014
  • Autophagy, the lysosomal degradation pathway, is an intracellular recycling system that is necessary for the metabolic benefits of exercise and for producing lasting beneficial effects of exercise in various diseases. However, the most recent studies have only examined the effect of a single bout of exercise or resistance exercise on autophagic responses. To determine the differential effects of acute and chronic exercise on the expression of autophagy-related genes in D. melanogaster, white-eyed mutant D. melanogaster were assigned randomly to four groups: control, acute exercise, 2 hr chronic exercise, and 3 hr chronic exercise. The flies were exercised using a mechanized platform known as the Power Tower. Our results revealed that a single bout of exercise resulted in increased mRNA levels of the Atg8a gene (~20%, p<0.05). However, Atg1 and Atg6 mRNA expression were not induced by acute exercise. Transcript levels of Atg6 (~29%, p<0.05) related to the nucleation of autophagosomes were significantly induced by 2 hr of chronic exercise. However, this chronic exercise was not enough to increase Atg1 and Atg8a mRNA expression. On the other hand, 3 hr of exercise for 7 days significantly increased Atg1, Atg6, and Atg8a gene expression-about 57%, 37%, and 71%, respectively (p<0.05). These results suggest that a single bout of exercise is not enough to induce full activation of selected autophagy-related genes in D. melanogaster. Our results demonstrated that chronic regular exercise induced autophagy-related gene expression, suggesting that chronic regular exercise training might be required to activate autophagic responses important for producing beneficial effects of exercise in various diseases.

Expressional Analysis of Superoxide Dismutase in Olive Flounder (Paralichthys olivaceus) against Viral Hemorrhagic Septicemia Virus Infection (Viral hemorrhagic septicemia virus (VHSV) 감염에 대한 넙치 superoxide dismutase(Of-SOD)의 발현분석)

  • Lee, Young Mee;Kim, Jung-Eun;Noh, Jae Koo;Kim, Hyun Chul;Park, Choul-Ji;Park, Jong-Won;Kim, Kyung-Kil;Lee, Jeong-Ho
    • Journal of Life Science
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    • v.24 no.12
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    • pp.1371-1377
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    • 2014
  • Superoxide dismutase is a family of important antioxidant metalloenzymes and catalyzes the dismutation of toxic superoxide anions into dioxygen and hydrogen peroxide. A recent study identified the partial superoxide dismutase (SOD) gene in olive flounder (Paralichthys olivaceus). The same study reported that it strongly induced benzo[a]pyrene and that it was an indicator of aquatic oxidative stress responses. However, its transcriptional response against viral infection has not been investigated. In the present study, the spatial and temporal expression profiles were analyzed to investigate the function of Of-SOD in the antiviral response. The Of-SOD transcripts were ubiquitously detected at various levels in diverse tissues in a real-time PCR. The expression of Of-SOD was significantly higher in the muscles, liver, and brain but extremely low in the stomach and spleen. Following a VHSV challenge, the expression of Of-SOD increased within 3 h in the kidneys and decreased to the original level 2 days postchallenge. In muscle, liver, and brain, Of-SOD mRNA was similarly up-regulated at 3-6 h postchallenge and then decreased to the basal level. Although the expression pattern and induction time differed slightly depending on the tissue, the transcript of Of-SOD consistently increased in the acute infection response, but the expression was low in the chronic response. The expression of Of-SOD was induced after the VHSV infection, and Of-SOD was probably involved in the immune response against the viral challenge. These results suggest that SOD may play important roles in the immune defense system of P. olivaceus and perhaps contribute to the protective effects against oxidative stress in olive flounder.

Context Sharing Framework Based on Time Dependent Metadata for Social News Service (소셜 뉴스를 위한 시간 종속적인 메타데이터 기반의 컨텍스트 공유 프레임워크)

  • Ga, Myung-Hyun;Oh, Kyeong-Jin;Hong, Myung-Duk;Jo, Geun-Sik
    • Journal of Intelligence and Information Systems
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    • v.19 no.4
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    • pp.39-53
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    • 2013
  • The emergence of the internet technology and SNS has increased the information flow and has changed the way people to communicate from one-way to two-way communication. Users not only consume and share the information, they also can create and share it among their friends across the social network service. It also changes the Social Media behavior to become one of the most important communication tools which also includes Social TV. Social TV is a form which people can watch a TV program and at the same share any information or its content with friends through Social media. Social News is getting popular and also known as a Participatory Social Media. It creates influences on user interest through Internet to represent society issues and creates news credibility based on user's reputation. However, the conventional platforms in news services only focus on the news recommendation domain. Recent development in SNS has changed this landscape to allow user to share and disseminate the news. Conventional platform does not provide any special way for news to be share. Currently, Social News Service only allows user to access the entire news. Nonetheless, they cannot access partial of the contents which related to users interest. For example user only have interested to a partial of the news and share the content, it is still hard for them to do so. In worst cases users might understand the news in different context. To solve this, Social News Service must provide a method to provide additional information. For example, Yovisto known as an academic video searching service provided time dependent metadata from the video. User can search and watch partial of video content according to time dependent metadata. They also can share content with a friend in social media. Yovisto applies a method to divide or synchronize a video based whenever the slides presentation is changed to another page. However, we are not able to employs this method on news video since the news video is not incorporating with any power point slides presentation. Segmentation method is required to separate the news video and to creating time dependent metadata. In this work, In this paper, a time dependent metadata-based framework is proposed to segment news contents and to provide time dependent metadata so that user can use context information to communicate with their friends. The transcript of the news is divided by using the proposed story segmentation method. We provide a tag to represent the entire content of the news. And provide the sub tag to indicate the segmented news which includes the starting time of the news. The time dependent metadata helps user to track the news information. It also allows them to leave a comment on each segment of the news. User also may share the news based on time metadata as segmented news or as a whole. Therefore, it helps the user to understand the shared news. To demonstrate the performance, we evaluate the story segmentation accuracy and also the tag generation. For this purpose, we measured accuracy of the story segmentation through semantic similarity and compared to the benchmark algorithm. Experimental results show that the proposed method outperforms benchmark algorithms in terms of the accuracy of story segmentation. It is important to note that sub tag accuracy is the most important as a part of the proposed framework to share the specific news context with others. To extract a more accurate sub tags, we have created stop word list that is not related to the content of the news such as name of the anchor or reporter. And we applied to framework. We have analyzed the accuracy of tags and sub tags which represent the context of news. From the analysis, it seems that proposed framework is helpful to users for sharing their opinions with context information in Social media and Social news.