• The Plant Pathology Journal
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• 제29권4호
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• pp.454-459
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• 2013

The Potexvirus Alternanthera mosaic virus (AltMV) has multifunctional triple gene block (TGB) proteins, among which our studies have focused on the properties of the TGB1 protein. The TGB1 of AltMV has functions including RNA binding, RNA silencing suppression, and cell-to-cell movement, and is known to form homologous interactions. The helicase domains of AltMV TGB1 were separately mutated to identify which regions are involved in homologous TGB1 interactions. The yeast two hybrid system and Bimolecular Fluorescence Complementation (BiFC) in planta were utilized to examine homologous interactions of the mutants. Helicase motif I of AltMV TGB1 was found to be critical to maintain homologous interactions. Mutations in the remaining helicase motifs did not inhibit TGB1 homologous interactions. In the absence of homologous interaction of TGB1, subcellular localization of helicase domain I mutants showed distinctively different patterns from that of WT TGB1. These results provide important information to study viral movement and replication of AltMV.

• Asian Pacific Journal of Cancer Prevention
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• 제15권14호
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• pp.5741-5745
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• 2014

Purpose: To study the expression of insulin-like growth factor binding proteins (IGFBPs) in paclitaxel-treated gastric cancer SGC-7901 cells, and to further investigate underlying mechanisms. Materials and Methods: Real time PCR and Western blot assays were applied to detect the mRNA and protein expression of IGFBP-2, -3 and -5 after paclitaxel (10 nM) treatment of SGC-7901 cells. In addition IGFBP-3 expression was silenced by RNA interference to determine effects. Cell viability was determined by MTT assay. Cell cycling and apoptosis were assessed by flow cytometry. Results: Compared to the control group, only IGFBP-3 expression was elevated significantly after paclitaxel (10 nM) treatment (p<0.05). Paclitaxel treatment caused cell cycle arrest and apoptosis via downregulating Bcl-2 expression. However, the effect could be abrogated by IGFBP-3 silencing. Conclusions: IGFBP-3 exhibits anti-apoptotic effects on paclitaxel-treated SGC-7901 cells via elevating Bcl-2 expression.

• 한국수정란이식학회지
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• 제28권1호
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• pp.57-61
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• 2013

Microtubule-associated protein 1B (MAP1B), a member of MAP1 family, plays a key role in neuronal development. MAP1B binds to many kinds of proteins directly or indirectly. This study was performed to investigate whether MAP1B interacts with GAPDH in bovine follicles using immunoprecipitation (IP) with Western blot analysis and immunohistochemisty. The mRNA expressions of MAP1B and glyceraldehydes 3-phosphate dehydrogenase (GAPDH) were down-regulated in bovine follicular cystic follicles (FCF). In parallel with the mRNA levels, their protein levels were also down-regulated in FCFs. In addition, MAP1B and GAPDH were co-localized at the cytoplasm of follicles. IP with Western blot analysis showed that MAP1B bound to GAPDH in normal follicles, but their binding was absent in FCFs, suggesting a low level of MAP1B and/or GAPDH expressions in FCFs. Taken together, these results suggest that MAP1B interacted with GAPDH may play a role in bovine follicle development, and that GAPDH does not function always as a loading control in bovine follicles.

• 대한의생명과학회지
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• 제18권3호
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• pp.227-236
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• 2012

Previous study showed that swimming improved obesity but was not through $PPAR{\alpha}$ activation in liver and skeletal muscle in high fat diet-fed female mice with functioning ovaries as an animal model of obese premenopausal women. Thus, this study was aimed at investigation of the effects of swimming on the promotion of health and its molecular mechanism in adipose tissue of high fat diet-fed female mice. Eight-week-old female C57BL/6J mice were randomly divided into two groups (a non-swim control group and a swim group, n=8/group). Mice in the swim group swam for 2 h daily for 6 weeks in water bath with temperature of $35{\pm}1^{\circ}C$. All the animals received high fat diet (45% kcal fat) for 6 weeks. Reverse transcription-polymerase chain reaction was used to elucidate the molecular mechanism. Female mice subjected to swimming had significantly decreased body weight gain and white adipose tissue mass compared with the female control mice. Histological studies illustrated that swimming decreases the hepatic lipid accumulation. As expected, swimming did not affect the expression of mRNA levels of peroxisome proliferator-activated receptor (PPAR) ${\alpha}$ and $PPAR{\alpha}$ target genes responsible for mitochondrial fatty acid ${\beta}$-oxidation, such as carnitine palmitoyltransgerase-1 and medium chain acyl-CoA dehydrogenase in the white adipose tissue. However, mice that underwent 6-weeks of swimming exercise had decreased the mRNA expression of lipogenic genes, such as sterol regulatory element-binding proteins-1C and fatty acid synthase in comparison to sedentary control mice, with decreased $PPAR{\gamma}$ target genes involved in adipocyte-specific marker genes, such as adipocyte fatty acid binding protein and leptin in the white adipose tissue. These results suggest that swimming can effectively prevent obesity induced by high fat diet-fed, in part through down-regulation of adipogenesis and lipogenesis in white adipose tissue of female obese mice. Moreover, these results suggest that swimming maybe contributing the promotion of health through regulation of adipogenesis and lipogenesis in overweight premenopausal women.

• Molecules and Cells
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• 제39권8호
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• pp.625-630
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• 2016

The RNA-binding protein Rbfox3 is a well-known splicing regulator that is used as a marker for post-mitotic neurons in various vertebrate species. Although recent studies indicate a variable expression of Rbfox3 in non-neuronal tissues, including lung tissue, its cellular function in lung cancer remains largely unknown. Here, we report that the number of RBFOX3-positive cells in tumorous lung tissue is lower than that in normal lung tissue. As the transforming growth factor-${\beta}$ (TGF-${\beta}$) signaling pathway is important in cancer progression, we investigated its role in RBFOX3 expression in A549 lung adenocarcinoma cells. TGF-${\beta}1$ treatment inhibited RBFOX3 expression at the transcriptional level. Further, RBFOX3 depletion led to a change in the expression levels of a subset of proteins related to epithelial-mesenchymal transition (EMT), such as E-cadherin and Claudin-1, during TGF-${\beta}1$-induced EMT. In immunofluorescence microscopic analysis, mesenchymal morphology was more prominent in RBFOX3-depleted cells than in control cells. These findings show that TGF-${\beta}$-induced RBFOX3 inhibition plays an important role in EMT and propose a novel role for RBFOX3 in cancer progression.

• Molecules and Cells
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• 제38권5호
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• pp.416-425
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• 2015

NF-E2-related factor 2 (Nrf2), a basic leucine zipper transcription factor, has recently received a great deal of attention as an important molecule that enhances antioxidative defenses and induces resistance to chemotherapy or radiotherapy. In this study, we investigated the apoptosis-inducing and Nrf2- upregulating effects of quercetin on malignant mesothelioma (MM) MSTO-211H and H2452 cells. Quercetin treatment inhibited cell growth and led to upregulation of Nrf2 at both the mRNA and protein levels without altering the ubiquitination and extending the half-life of the Nrf2 protein. Following treatment with quercetin, analyses of the nuclear level of Nrf2, Nrf2 antioxidant response element-binding assay, Nrf2 promoter-luc assay, and RT-PCR toward the Nrf2-regulated gene, heme oxygenase-1, demonstrated that the induced Nrf2 is transcriptionally active. Knockdown of Nrf2 expression with siRNA enhanced cytotoxicity due to the induction of apoptosis, as evidenced by an increase in the level of proapoptotic Bax, a decrease in the level of antiapoptotic Bcl-2 with enhanced cleavage of caspase-3 and PARP proteins, the appearance of a sub-$G_0/G_1$ peak in the flow cytometric assay, and increased percentage of apoptotic propensities in the annexin V binding assay. Effective reversal of apoptosis was observed following pretreatment with the pan-caspase inhibitor Z-VAD. Moreover, Nrf2 knockdown exhibited increased sensitivity to the anticancer drug, cisplatin, presumably by potentiating the oxidative stress induced by cisplatin. Collectively, our data demonstrate the importance of Nrf2 in cytoprotection, survival, and drug resistance with implications for the potential significance of targeting Nrf2 as a promising strategy for overcoming resistance to chemotherapeutics in MM.

• 한국자원식물학회지
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• 제28권5호
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• pp.582-590
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• 2015

본 연구는 HRL의 3T3-L1 지방전구세포의 분화과정 중에 HRL이 지방의 축적에 미치는 영향을 확인하였다. MTT assay를 이용하여 세포 독성을 측정한 결과 100 ㎍/㎖의 농도에서도 세포증식에 영향을 미치지 않는 것을 확인하였고, 이와 같은 결과를 토대로 Oil Red O 염색법을 이용하여 지방세포 분화 억제능을 측정하였다. 그 결과, HRL의 경우 100 ㎍/㎖의 농도에서 82.25% 지방 축적 억제능을 나타내었다. 지방생성에 영향을 미치는 유전자 발현량을 측정하기 위해 RT-PCR법과 western blot법을 시행하였다. HRL은 SREBP-1c, PPARγ와 C/EBPα의 mRNA 발현을 억제시켰고, 지방생성에 영향을 미치는 효소인 FAS의 생성을 조절하는 것으로 나타났다. 또한, HRL 처리로 AMPKα의 단백질 발현이 증가하였으며, PPARγ의 발현량이 감소하는 것을 확인하였다. 이상의 결과들로부터 HRL은 AMPKα의 활성화를 통한 지방 합성을 억제를 보유하고 있는 바, 향후 항비만 기능성 소재로 활용될 수 있을 것으로 생각한다.

• 한국산학기술학회논문지
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• 제9권6호
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• pp.1795-1799
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• 2008

hnRNP K 단백질은 hnRNP 복합체를 구성하는 핵단백질들 중의 하나이며 시토신이 많은 RNA/DNA sequence에 잘 결합한다. 이 단백질은 핵 내에서만 머무르지 않고 핵과 세포질을 왕복하는 특징을 지니고 있다. hnRNP K의 기능을 조사하기 위하여 우선 hnRNP K와 상호 결합하는 세포내 단백질을 찾아내고자 하였다. 이를 위하여 본 연구에서는 이스트 two-hybrid 시스템을 사용하여 HeLa CDNA librar를 탐색하였다. 그 결과 얻은 클론들 중에는 사람의 hnRNP E1 (poly(rC) binding protein 1) cDNA (GenBank accession number XM_031585) 클론이 포함되어 있었다. 본 논문에서는 이스트 two-hybrid 시스템과 in vitro에서의 생화학적 실험을 통하여 hnRNP E1은 hnRNP K와 특이적으로 상호 결합한다는 것을 밝혔다.

• 생명과학회지
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• 제26권4호
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• pp.490-495
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• 2016

Brefeldin A (BFA)는 Eupenicillium brefeldianum에서 분리한 lactone계열의 항생제이며 ER에서 Golgi로 단백질 이송/전달을 억제히는 기능이 있다. 따라서 BFA를 세포에 처리 시 Golgi 기능 장애와 ER에서 단백질의 폴딩/조립의 문제로 인하여 ER에 기능 장애가 발생하는데 이를 소포체 스트레스(ER stress)라고 한다. 본 연구에서는 정상 astrocyte 세포와 C6 glioma 세포에서의 BFA처리에 따라 ER stress marker 단백질인 CHOP 발현 차이를 확인하였다. BFA 처리 시 CHOP 발현이 정상 astrocyte 세포에서 C6 glioma 세포에 비해 현저히 낮은 발현을 확인하였다. 하지만 CHOP mRNA 발현에서는 astrocyte 세포에서 발현 됨을 RT-PCR로 확인하였다. C6 glioma 세포와 비교하여 astrocyte 세포에서 BFA유도의 CHOP 단백질 발현이 낮은 원인은 proteasome 활성이 높음으로 기인됨을 proteasome inhibitor 실험과 proteasome 활성 측정을 통하여 확인하였다.

• Korean Circulation Journal
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• 제53권6호
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• pp.387-403
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• 2023

Background and Objectives: Myocardial ischemia and reperfusion injury (MIRI) has high morbidity and mortality worldwide. We aimed to explore the role of long noncoding RNA lysyl oxidase like 1 antisense RNA 1 (LOXL1-AS1) in cardiomyocyte pyroptosis. Methods: Hypoxia/reoxygenation (H/R) injury was constructed in human cardiomyocyte (HCM). The level of LOXL1-AS1, miR-761, phosphatase and tensin homolog (PTEN) and pyroptosis-related proteins was monitored by quantitative real-time polymerase chain reaction or western blot. Flow cytometry examined the pyroptosis level. Lactate dehydrogenase (LDH), creatine kinase-MB and cardiac troponin I levels were detected by test kits. Enzyme-linked immunosorbent assay measured the release of inflammatory cytokines. Dual-luciferase assay validated the binding relationship among LOXL1-AS1, miR-761, and PTEN. Finally, ischemia/reperfusion (I/R) animal model was constructed. Hematoxylin and eosin staining assessed morphological changes of myocardial tissue. NOD-like receptor pyrin domain-containing protein 3 (NLRP3) and casepase-1 expression was determined by immunohistochemistry. Results: After H/R treatment, LOXL1-AS1 and PTEN were highly expressed but miR-761 level was suppressed. LOXL1-AS1 inhibition or miR-761 overexpression increased cell viability, blocked the release of LDH and inflammatory cytokines (interleukin [IL]-1β, IL-18), inhibited pyroptosis level, and downregulated pyroptosis-related proteins (ASC, cleaved caspase-1, gasdermin D-N, NLRP3, IL-1β, and IL-18) levels in HCMs. LOXL1-AS1 sponged miR-761 to up-regulate PTEN. Knockdown of miR-761 reversed the effect of LOXL1-AS1 down regulation on H/R induced HCM pyroptosis. LOXL1-AS1 aggravated the MIRI by regulating miR-761/PTEN axis in vivo. Conclusions: LOXL1-AS1 targeted miR-761 to regulate PTEN expression, then enhance cardiomyocyte pyroptosis, providing a new alternative target for the treatment of MIRI.