• Bulletin of the Korean Chemical Society
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• 제28권1호
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• pp.77-80
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• 2007

As neurofilament proteins are major cytoskeletal components of neuron, abnormality of neurofilament is proposed in brain with neurodegenerative disorders such as Parkinson's disease (PD). Since oxidative stress might play a critical role in altering normal brain proteins, we investigated the oxidative modification of neurofilament-L (NF-L) induced by the reaction of cytochrome c with H2O2. When NF-L was incubated with cytochrome c and H2O2, the protein aggregation was increased in cytochrome c and H2O2 concentrationsdependent manner. Radical scavengers, azide, formate and N-acetyl cysteine, prevented the aggregation of NFL induced by the cytochrome c/H2O2 system. The formations of carbonyl group and dityrosine were obtained in cytochrome c/H2O2-mediated NF-L aggregates. Iron specific chelator, desferoxamine, prevented the cytochrome c/H2O2 system-mediated NF-L aggregation. These results suggest that the cytochrome c/H2O2 system may be related to abnormal aggregation of NF-L which may be involved in the pathogenesis of PD and related disorders.

• Clinical and Experimental Reproductive Medicine
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• 제33권3호
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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.

• Asian-Australasian Journal of Animal Sciences
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• 제24권1호
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• pp.110-124
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• 2011

Anatomically separate fat depots differ in size, function, and contribution to pathological states such as the metabolic syndrome. We isolated pre-adipocytes from different adipose depots, omental, subcutaneous and intramuscular, of beef cattle, and cultured in vitro to determine the basis for the variations and attribute these variations to the inherent properties of adipocyte progenitors. The proliferating cells from all depots before the confluence were harvested and the proteome was analyzed by a functional proteomic approach, involving 2-DE and MALDI-TOF/TOF. More than 252 protein spots were identified, selected and analyzed by Image Master (ver 7.0) and MALDI-TOF/TOF. Further, our analysis showed that there were specific differences in proteome expression patterns among proliferating precursor cells from the three depots. Sixteen proteins were found to be differentially expressed and these were identified as proteins involved in cellular processes, heat shock/chaperones, redox proteins, cytoskeletal proteins and metabolic enzymes. The results also enabled us to understand the basic roles of these proteins in different inherent properties exhibited by adipose tissue depots.

• Biomedical Science Letters
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• 제10권3호
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• pp.179-186
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• 2004

We examined a role of proteasome, the non-lysosomal multicatalytic protease complex,on the differentiation of chick embryonic myoblasts in culture. The peptide hydrolyzing activities of proteasome were found to change; the hydrolyzing activity against N-succinyl-Leu-Leu- Val- Tyr-7 -amido-4-methy1coumarin (SLLVY-AMC) was prominent and increased with myogenic differentiation. Proteasome inhibitors, N-carbobenzoxy-Leu-Leu-norvalinal (MG115) and N-carbobenzoxy-Ile-Glu (O-t-butyl)-Ala-Leucinal (PSI), blocked membrane fusion of myoblasts as well as the SLLVY-AMC hydrolyzing activity. Those inhibitory activities of the agents occurred in parallel, but were reversible and both cell fusion and the peptidase activity were restored when the agents were withdrawn from the culture medium. On the other hand, the agents caused accumulation of the ubiquitinylated proteins in the cytoskeletal proteins. These results suggest that each of the peptide hydrolyzing activities of proteasome is independently regulated during the myogenic differentiation and the chymotrypsin-like activity may play an important role in that process.

• Journal of Microbiology and Biotechnology
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• 제21권12호
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• pp.1211-1227
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• 2011

C57BL/6J mice have been widely used as a diet-induced obesity model because they trigger common features of the human metabolic syndrome. In the present study, C57BL/6J male mice were fed either a high-fat diet (HFD) or normal diet (ND) during a 24-week period, and then the age-dependent liver proteome of mice in two groups was analyzed using 2-DE combined with MALDI-TOF-MS. Among identified proteins, up-regulated proteins were subdivided to early (during the first 4 weeks) and late (20~24 weeks) markers that played a role in diet-induced obesity development. Important early markers included ketohexokinase and prohibitin, and late markers included the 75 kDa glucose-regulated protein, citrate synthase, and selenium-binding liver protein. Of these, the 75 kDa glucosere-gulated protein has already been linked to obesity; however, prohibitin protein involved in obesity was identified for the first time in this study. In order to validate the proteomic results and gain insight into metabolic changes between the two groups, we further confirmed the expression pattern of some proteins of interest by Western blot analysis. Combined results of proteomic analysis with Western blot analysis revealed that antioxidant enzymes were progressively decreased, whereas cytoskeletal proteins were time-dependently increased in HFD mice.

• Genomics & Informatics
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• 제21권3호
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• pp.36.1-36.19
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• 2023

The LIM domain-containing proteins are dominantly found in plants and play a significant role in various biological processes such as gene transcription as well as actin cytoskeletal organization. Nevertheless, genome-wide identification as well as functional analysis of the LIM gene family have not yet been reported in the economically important plant sorghum (Sorghum bicolor L.). Therefore, we conducted an in silico identification and characterization of LIM genes in S. bicolor genome using integrated bioinformatics approaches. Based on phylogenetic tree analysis and conserved domain, we identified five LIM genes in S. bicolor (SbLIM) genome corresponding to Arabidopsis LIM (AtLIM) genes. The conserved domain, motif as well as gene structure analyses of the SbLIM gene family showed the similarity within the SbLIM and AtLIM members. The gene ontology (GO) enrichment study revealed that the candidate LIM genes are directly involved in cytoskeletal organization and various other important biological as well as molecular pathways. Some important families of regulating transcription factors such as ERF, MYB, WRKY, NAC, bZIP, C2H2, Dof, and G2-like were detected by analyzing their interaction network with identified SbLIM genes. The cis-acting regulatory elements related to predicted SbLIM genes were identified as responsive to light, hormones, stress, and other functions. The present study will provide valuable useful information about LIM genes in sorghum which would pave the way for the future study of functional pathways of candidate SbLIM genes as well as their regulatory factors in wet-lab experiments.

• BMB Reports
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• 제43권10호
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• pp.656-663
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• 2010

Amyloid precursor protein (APP), which is critically involved in the pathogenesis of Alzheimer's disease (AD), is cleaved by gamma/epsilon-secretase activity and results in the generation of different lengths of the APP Intracellular C-terminal Domain (AICD). In spite of its small size and short half-life, AICD has become the focus of studies on AD pathogenesis. Recently, it was demonstrated that AICD binds to different intracellular binding partners ('adaptor protein'), which regulate its stability and cellular localization. In terms of choice of adaptor protein, phosphorylation seems to play an important role. AICD and its various adaptor proteins are thought to take part in various cellular events, including regulation of gene transcription, apoptosis, calcium signaling, growth factor, and $NF-{\kappa}B$ pathway activation, as well as the production, trafficking, and processing of APP, and the modulation of cytoskeletal dynamics. This review discusses the possible roles of AICD in the pathogenesis of neurodegenerative diseases including AD.

• Proceedings of the KSME Conference
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• 대한기계학회 2008년도 추계학술대회A
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• pp.1455-1457
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• 2008

Cell adhesion is a coordinated process involving initial binding of integrin receptors to extracellular matrix (ECM), recruitment of adhesion proteins, and focal adhesion assembly. The formation of mechanically stable focal adhesion assembly of cells within surrounding ECM is a key parameter to direct numerous cellular functions including cell migration, differentiation, and apotosis. With current cell adhesion assays, it is difficult to understand contributions of each coordinated event on evolution of cell adhesion strengthening since cells spontaneously spread upon their adhesion to the substrate, thus remodeling their cytoskeletal structure. In this presentation, novel approaches for analysis of cell adhesion strengthening process based on the combination of mechanical device, micro-patterned substrates, and molecular biological techniques will be discussed.

• Clinical and Experimental Reproductive Medicine
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• 제38권1호
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• pp.1-5
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• 2011

The maturation process of mammalian oocytes accompanies an extensive rearrangement of the cytoskeleton and associated proteins. As this process requires a delicate interplay between the cytoskeleton and its regulators, it is often targeted by various external and internal adversaries that affect the congression and/or segregation of chromosomes. Asymmetric cell division in oocytes also requires specific regulators of the cytoskeleton, including formin-2 and small GTPases. Recent literature providing clues regarding how actin filaments and microtubules interact during spindle migration in mouse oocytes are highlighted in this review.

• Journal of Life Science
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• 제24권12호
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• pp.1276-1283
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• 2014

Cell adhesion molecules determine the cell-cell binding and the interactions between cells and extracellular signals. Cell-cell junctional complexes, which maintain the structural integrity of tissues, consist of more than 50 proteins including multi-PDZ domain protein 1 (MUPP1). MUPP1 contains 13 postsynaptic density-95/disks large/zonula occludens-1 (PDZ) domains and serves a scaffolding function for transmembrane proteins and cytoskeletal proteins or signaling proteins, but the mechanism how MUPP1 links and stabilizes the juxtamembrane proteins has not yet been elucidated. We used the yeast two-hybrid system to identify proteins that interact with PDZ domains of MUPP1. We found an interaction between MUPP1 and cell adhesion molecule 1 (Cadm1, also known as SynCAM1, Necl-2, or TSLC1). Cadm1 bound to the second PDZ domain of MUPP1. The carboxyl (C)-terminal end of Cadm1 has a type II PDZ-association motif (-Y-F-I) which was essential for the interaction with MUPP1 in the yeast two-hybrid assay. MUPP1 also bound to the C-terminal cytoplasmic tail region of other Cadm family members (Cadm2, Cadm3, and Cadm4). In addition, these protein-protein interactions were observed in the glutathione S-transferase (GST) pull-down assay and by co-immunoprecipitation. Anti-MUPP1 antibody co-immunoprecipitated Cadm1 and Cadm4 with MUPP1 from mouse brain extracts. These results suggest that MUPP1 could mediate interaction between Cadms and cytoskeletal proteins.