• Journal of Life Science
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• v.26 no.3
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• pp.282-288
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• 2016

Protein-protein interactions regulate the subcellular localization and function of receptors, enzymes, and cytoskeletal proteins. Proteins containing the postsynaptic density-95/disks large/zonula occludens-1 (PDZ) domain have potential to act as scaffolding proteins and play a pivotal role in various processes, such as synaptic plasticity, neural guidance, and development, as well as in the pathophysiology of many diseases. Multi-PDZ domain protein 1 (MUPP1), which has 13 PDZ domains, has a scaffolding function in the clustering of surface receptors, organization of signaling complexes, and coordination of cytoskeletal dynamics. However, the cellular function of MUPP1 has not been fully elucidated. In the present study, a yeast two-hybrid system was used to identify proteins that interacted with the N-terminal PDZ domain of MUPP1. The results revealed an interaction between MUPP1 and Wdpcp (formerly known as Fritz). Wdpcp was identified as a planar cell polarity (PCP) effector, which is known to have a role in collective cell migration and cilia formation. Wdpcp bound to the PDZ1 domain but not to other PDZ domains of MUPP1. The C-terminal end of Wdpcp was essential for the interaction with MUPP1 in the yeast two-hybrid assay. This interaction was further confirmed in a glutathione S-transferase (GST) pull-down assay. When coexpressed in HEK-293T cells, Wdpcp was coimmunoprecipitated with MUPP1. In addition, MUPP1 colocalized with Wdpcp at the same subcellular region in cells. Collectively, these results suggest that the MUPP1-Wdpcp interaction could modulate actin cytoskeleton dynamics and polarized cell migration.

• Tuberculosis and Respiratory Diseases
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• v.55 no.1
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• pp.21-30
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• 2003

Background : Mucin synthesis in airways has been reported to be regulated by the epidermal growth factor receptor (EGFR) system. Epidermal growth factor receptor transactivation was identified as a critical element in G-protein-coupled receptors (GPCRs)-induced mitogenic signaling. EGF receptor transactivation by G-protein-coupled receptors requires metalloproteinase cleavage of proHB-EGF. This study was hypothesized that lipopolysaccharide (LPS)-induced mucin production associates with epidermal growth factor receptor transactivation, and MUC5AC production associates with epidermal growth factor receptor transactivation by G-protein-coupled receptors that regulates by metalloproteinase. Method : MUC5AC mucin production was examined in NCI-H292 cells and MUC5AC protein synthesis was assessed using ELISA. For the evaluation of mechanism of LPS-induced MUC5AC production, $TNF{\alpha}$ was measured using ELISA with or without pretreatment of heterotrimeric G-protein inhibitor, mastoparan. MUC5AC protein was measure with pretreatment of polyclonal $TNF{\alpha}$ antibody or mastoparan on LPS-induced MUC5AC production. For the evaluation of relation of G-protein and MUC5AC production, G-protein stimulant, mastopara-7, or matrix metalloproteinase, ADAM10, was added to NCI-H292 cells. MUC5AC protein was measure with pretreatment of polyclonal EGF antibody on mastoparan-7-induced MUC5AC production. Results : LPS alone did not increase significantly MUC5AC production. LPS with $TNF{\alpha}$ induced dose-dependently MUC5AC production in NCI-H292 cells. LPS increased dose-dependently $TNF{\alpha}$ secretion, which was inhibited by mastoparan. LPS with $TNF{\alpha}$-induced MUC5AC production was inhibited by neutralizing polyclonal $TNF{\alpha}$ antibody, mastoparan or AG 1472. Mastoparan-7 or ADAM10 increased dose-dependently MUC5AC production, which was inhibited by polyclonal neutralizing EGF antibody. Conclusion : In LPS-induced MUC5AC synthesis, LPS causes $TNF{\alpha}$ secretion, which induces EGFR expression. EGFR tyrosine kinase phosphorylation result in MUC5AC production. EGF-R transactivation by G-protein-coupled receptors requires matrix metalloproteinase cleavage of proHB-EGF.

• The Korean Journal of Food And Nutrition
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• v.26 no.4
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• pp.806-813
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• 2013

Agrimonia pilosa Ledeb. is a medicinal plant with anti-tumor, anti-oxidant, anti-inflammatory and anti-hyperglycemic activities. However, few studies of the anti-diabetic effect of A. pilosa on insulin resistance status have been performed. In the present study, the anti-diabetic effect of A. pilosa water extract (AP) was determined by investigating its ${\alpha}$-glucosidase inhibitory property, glucose utilization, and uptake, as well as insulin resistance mechanism of action in C2C12 skeletal muscle cells. Compared to positive control (acarbose), AP ($10mg/m{\ell}$) showed a similar ${\alpha}$-glucosidase inhibitory capacity. Glucose uptake was significantly increased by $1{\mu}m$ insulin treatment (p<0.05). However, palmitic acid (FFA, 1 mM) induced muscle insulin resistance and glucose uptake dysfunction. On the other hand, AP ($10{\mu}g/m{\ell}$) was capable of reversing the FFA-induced insulin resistance in C2C12 myotubes. Compared to control, AP ($100{\mu}g/m{\ell}$ without insulin) significantly increased the utilization of glucose (p<0.05) in C2Cl2 myotubes cultured in normal glucose (7 mM). AP treatment significantly increased the relative mRNA and protein expression levels of Akt. In particular, the effect of A. pilosa on the insulin signaling system is associated with the up-regulation of Akt genes and glucose uptake in C2Cl2 myotubes. These results suggest that A. pilosa is useful in the prevention of diabetes and the treatment of hyperglycemic disorders.

• Journal of Yeungnam Medical Science
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• v.24 no.1
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• pp.24-40
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• 2007

Background : Accumulating evidence shows that interleukin(IL)-1 plays a critical role in inflammation and connective tissue destruction observed in both osteoarthritis and rheumatoid arthritis. IL-1 induces gene expression related to cytokines, chemokines and matrix metalloproteinases by activation of many different transcription factors. Materials and Methods : The chondrosarcoma cell line, SW1353, is known to be a valuable in vitro system for investigating catabolic gene regulation by IL-$1{\beta}$ in chondrocytic cells. To explore and analyze the changes in gene expression by IL-1 responsible for arthritis, SW1353 was treated with IL-1 for 1, 6 and 24 h and then total RNAs were purified for each time. The changes in gene expression were analyzed with 17k human cDNA microarrays and validated by semi-quantitative RT-PCR. Results : Greater than a two-fold change was observed in 1,200 genes including metallothioneins, matrix metalloproteinases, extracellular matrix proteins, antioxidant proteins, cytoskeleton proteins, cell cycle regulatory proteins, proteins for cell growth and apoptosis, signaling proteins and transcription factors. These changes appeared to be correlate with the pathophysiological changes observed in early osteoarthritis. Conclusion : cDNA microarray analysis revealed a marked variability in gene expression, and provided insight into the overall molecular changes. The result of this study provide initial information for further studies to identify therapeutic targets in osteoarthritis pathogenesis.

• Journal of Life Science
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• v.24 no.12
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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.

• Development and Reproduction
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• v.6 no.1
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• pp.55-66
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• 2002

Embryonic stem cells(ES cells) are derived from the inner cell mass(ICM) of blastocysts, which have the potentials to remain undifferentiated, to proliferate indefinitely in vitro, to differentiate into the derivates of three embryonic germ layers. ES cells are an attractive model system for studying the initial developmental decisions and their molecular mechanisms during embryogenesis. Additionally, ES cells of significant interest to those characterizing the various gene functions utilizing transgenic and gene targeting techniques. We investigated the effects of reproductive hormones, gonadotropins(GTH) and steroids on the induction of differentiation and expressions of their receptor genes using the newly established mouse ES cells. We collected the matured blastocysts of inbred mice C57BL/6J after superovulation and co-cultured with mitotically inactivated STO feeder cells. After 5 passages, we confirmed the expression alkaline phosphatase(Alk P) activity and SSEA-1, 3, 4 expressions. The protocol devised for inducing ES differentiation consisted of an aggregation steps, after 5 days as EBs in hormone treatments(FSH, LH, E$_2$, P$_4$, T) that allows complex signaling to occur between the cells and a dissociation step, induced differentiation through attachment culture during 7 days in hormone treatments. Hormone receptors were not increased in dose-dependent manner. All hormone receptors in ES cells treated reproductive hormones were expressed lower than those of undifferentiated ES cell except for LHR expression in E$_2$-treated ES cells group. After hormone induced differentiation, at least some of the cells are not terminally differentiated, as is evident from the expression of Oct-4, a marker of undifferentiated. To assess their differentiation by gene expression, we analyzed the expression of 7 tissue-specific markers from all three germ layers. Most of hormone-treated group increased in the expression of gata-4 and $\alpha$ -fetoprotein, suggesting reproductive hormone allowed or induced differentiation of endoderm.

• Journal of Life Science
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• v.25 no.5
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• pp.594-600
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• 2015

Protein-protein interactions have a critical role in the regulation of many cellular functions. Postsynaptic density-95/disks large/zonula occludens-1 (PDZ) domain is one of domains that mediate protein-protein interactions. PDZ domains typically bind to the specific motif at the carboxyl (C)-terminal end of partner proteins. Multi-PDZ domain protein 1 (MUPP1), which has 13 PDZ domains, serves a scaffolding function for structure proteins and signaling proteins, but the cellular function of MUPP1 has not been fully 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 muskelin. Muskelin was recently identified as a GABA_A receptor (GABA_AR) α1 subunit binding protein and known to have a role in receptor endocytosis and degradation. Muskelin bound to the 3^rd PDZ domain, but not to other PDZ domains of MUPP1. The C-terminal end of muskelin was essential for the interaction with MUPP1 in the yeast two-hybrid assay. When co-expressed in HEK-293T cells, muskelin but not the C-terminal deleted muskelin was co-immunoprecipitated with MUPP1. In addition, MUPP1 co-localized with muskelin at the same subcellular region in cells. These findings collectively suggest that MUPP1 or its interacting proteins could modulate GABA_AR trafficking and turnover through the interaction with muskelin.

• Korean Journal of Microbiology
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• v.46 no.3
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• pp.240-247
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• 2010

Pseudomonas aeruginosa is a Gram (-) opportunistic human pathogen causing a wide variety of infections on lung, urinary tract, eyes, and burn wound sites and quorum sensing (QS), a cell density-sensing mechanism plays an essential role in Pseudomonas pathogenesis. In order to investigate the importance of QS in the Pseudomonas infections of Korean patients, we isolated 189 clinical strains of P. aeruginosa from the patients in Pusan Paik Hospital, Busan, South Korea. The QS signal production of these clinical isolates was measured by signal diffusion assay on solid media using reporter strains. While most clinical strains (79.4%) produced the QS signals as similar level as a wild type strain, PAO1 did, where LasR, the initial QS signal sensor-regulator was fully activated, a minority of them (4.2%) produced much less QS signals at the level to which LasR failed to respond. Similarly, while 72.5% of the clinical isolates produced QS signals enough to activate QscR, an another QS signal sensor-regulator, some few of them (9%) produced the QS signals at much lower level where QscR was not activated. For further analysis, we selected 74 clinical strains that were obtained from the patients under suspicion of Pseudomonas infection and investigated the total protease activity that is considered important for virulence. Interestingly, significant portion of them showed very low protease activity (44.6%) or no detectable protease activity (12.2%). When the biofilm-forming ability that is considered very important in chronic infection was examined, most isolates showed lower biofilm-forming activity than PAO1. Similarly, significant portion of clinical isolates showed reduced motility (reduced swarming activity in 51.4% and reduced twitching activity in 41.9%), or non-detectable motility (swarming-negative in 28.4% and twitching-negative in 28.4%). Our result showed that the clinical isolates that produced QS signals at the similar level to wild type could have significantly reduced activities in the protease production, biofilm formation, and motility, and some clinical isolates had unique patterns of motility, biofilm formation, and protease production that are not correlated to their QS activity.

• Journal of Life Science
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• v.24 no.8
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• pp.820-826
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• 2014

The localization to specific subcellular sites and the regulation of cell surface receptors and channels are crucial for proper functioning. Postsynaptic density-95/Disks large/Zonula occludens-1 (PDZ)-domain is involved in recognition of and interaction between various proteins, by which the localization and the regulation are mediated. Multi-PDZ domain protein 1 (MUPP1) contains 13 PDZ domains. MUPP1 serves a scaffolding function for structure proteins and signaling proteins, but the mechanism how MUPP1 is stabilized and signalized 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 Parkin. Parkin is an E3 ubiquitin ligase. Loss-of-function mutations of Parkin gene are known to cause an autosomal recessive juvenile parkinsonism. Parkin bound to the $12^{th}$ PDZ domain, but not to other PDZ domains of MUPP1. The C-terminal end of Parkin has a type II PDZ-association motif, which was essential for the interaction with MUPP1 in the yeast two-hybrid assay. When co-expressed in HEK-293T cells, Parkin co-localized with MUPP1. When co-expressed with ubiquitin in HEK-293T cells, MUPP1 has been strongly ubiquitinated by Parkin. These findings collectively suggest that MUPP1 is a novel substrate of Parkin and its function or stability could be modulated by Parkin-mediated ubiquitination.

• Journal of Life Science
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• v.30 no.11
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• pp.999-1006
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• 2020

The suppression of neuroinflammatory responses in microglial cells, well known as the main immune cells in the central nervous system (CNS), are considered a key target for improving the progression of neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and Huntington's disease. Teleogryllus emma is widely consumed around the world for its broad-spectrum therapeutic effect. In a previous work, we performed transcriptome analysis on T. emma in order to obtain the diversity and activity of its antimicrobial peptides (AMPs). AMPs are found in a variety of species, from microorganisms to mammals. They have received much attention as candidates oftherapeutic drugs for the treatment of inflammation-associated diseases. In this study, we investigated the anti-neuroinflammatory effect of Teleogryllusine (VKWKRLNNNKVLQKIYFVKI-NH₂) derived from T. emma on lipopolysaccharide (LPS) induced BV-2 microglia cells. Teleogryllusine significantly inhibited nitric oxide (NO) production without cytotoxicity, and reducing pro-inflammatory enzymes expression such as inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). In addition, Telegryllusine also inhibited the expression of pro-inflammatory cytokines such as interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) through down-regulation of the mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NF-κB) signaling pathway. These results suggest that T. emma-derived Teleogryllusine could be a good source of functional substances that prevent neuroinflammation and neurodegenerative diseases.