Korea Research Institute of Bioscience and Biotechnology
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Wnt5a-induced docking of Plk1 on HEF1 promotes HEF1 translocation and tumorigenesis
No full textBackground: Upregulation of human enhancer filamentation 1 (HEF1/NEDD9/Cas-L) and Polo-like kinase 1 (Plk1) is closely correlated with metastasis of human cancer. However, the mechanism by which the overexpression of HEF1 or Plk1 stimulates cancer metastasis and induces tumorigenesis remains enigmatic. In addition, the accumulation of HEF1 at the focal adhesion (FA) is known to be an essential event in cancer cell migration, but the mechanism of how HEF1 is targeted to the FA remains yet to be unveiled.
Objective: This study was performed to elucidate the FA docking mechanism of HEF1 and to determine its effect on tumorigenesis.
Methods: To confirm the effect of the kinase on HEF1 translocation, various expression-knockdown stable cell lines were generated using a lentivirus system, and the effect of the HEF1-Plk1 complex on tumorigenesis was confirmed using a xenograft mouse model.
Results: Here, we show that Wnt5a-dependent Plk1 binding to HEF1 is critically required for HEF1 translocation to the FA. We also confirmed that Plk1 and CK1δ activities essential for HEF1 translocation are induced by Wnt5a. Finally, we confirmed the induction of tumorigenesis by the HEF1-Plk1 complex in the xenograft mouse model.
Conclusion: Our data collectively unveil the Wnt5a-CK1δ-HEF1-Plk1-FA remodeling pathway that governs HEF1 transportation to the FA to induce cell migration and tumorigenesis. This study sheds light on a mechanism underlying tumorigenesis and provides new strategies for anticancer therapy.
Pollen-mediated flow of bar gene in transgenic herbicide-resistant turf grass Zoysia japonica
No full textWeed control can be most effectively achieved through the use of herbicide-resistance transgene. A preliminary study bar-transgenic Zoysia japonica posed no serious risk on the unintended escape of the transgene from its cultivation site. The present follow-up investigation of the dispersion of pollen and its short-distance escape outside of the bar-transgenic Zoysia japonica habitats were ascertained in terms of environmental factors affecting anthesis and pollen viability. In a 24-h day cycle, zoysiagrass pollen was released predominantly between 08:00 and 10:00, and the pollen was most viable during the same time interval. Optimal temperature and humidity for pollen viability was 15？20 °C and 80？90%, respectively. The pollen germinated in 120 min after anthesis, but under cloudy conditions germination time doubled. No differences in pollen viability/longevity between the transgenic and non-transgenic plants were observed. The pollen-mediated gene flow of transgenic Zoysia japonica to wild-type non-transgenic zoysiagrass species was monitored by measuring the cross-over rate of the bar gene in the context of three different models. At distances within 5 m, the rate of gene flow ranged from 3 to 5.7% according to the models used. The greater the distance from the transgenic plant site, the lower the gene flow rate. The furthest transgene detected was 38 m away and exhibited a 0.25% gene flow rate. The radial model yielded a 3.7% escape rate within a 3 m radius and was wind direction dependent. The distance- and direction-dependent gene flow events were influenced by wind direction and velocity during flowering season.
Genetic characterization of feline parvovirus isolate Fe-P2 in Korean cat and serological evidence on its infection in wild leopard cat and Asian badger
No full textFeline parvovirus (FPV) is a small, non-enveloped, single-stranded DNA virus that infects cats. We recently isolated a feline parvovirus Fe？P2 strain from a dead stray cat in Iksan, 2017. Its partial genomic sequence (4,643 bases) was obtained, and phylogenetic analysis based on the VP2 nucleotide sequence showed that the FPV Fe-P2 strain was closely related to the FPV isolate Gigucheon in cat, 2017 (MN400978). In addition, we performed a serum neutralization (SN) test with the FPV isolates in various mammalian sera. These were from raccoon dog, water deer, Eurasian otter, Korean hare, leopard cat, and Asian badger, which were kindly provided by Chungnam Wild Animal Rescue Center. Notably, serological evidence of its infection was found in Asian badger, Meles leucurus (2/2) and leopard cat, Prionailurus bengalensis (5/8) through SN tests, whereas there was no evidence in raccoon dog, water deer, Eurasian otter, and Korean hare based on the collected sera in this study. These findings might provide partial evidence for the possible circulation of FPV or its related viruses among wild leopard cat and Asian badger in Korea. There should be additional study to confirm this through direct detection of FPVs in the related animal samples.
Mitophagy deficiency increases NLRP3 to induce brown fat dysfunction in mice
No full textAlthough macroautophagy/autophagy deficiency causes degenerative diseases, the deletion of essential autophagy genes in adipocytes paradoxically reduces body weight. Brown adipose tissue (BAT) plays an important role in body weight regulation and metabolic control. However, the key cellular mechanisms that maintain BAT function remain poorly understood. in this study, we showed that global or brown adipocyte-specific deletion of pink1, a Parkinson disease-related gene involved in selective mitochondrial autophagy (mitophagy), induced BAT dysfunction, and obesity-prone type in mice. Defective mitochondrial function is among the upstream signals that activate the NLRP3 inflammasome. NLRP3 was induced in brown adipocyte precursors (BAPs) from pink1 knockout (KO) mice. Unexpectedly, NLRP3 induction did not induce canonical inflammasome activity. Instead, NLRP3 induction led to the differentiation of pink1 KO BAPs into white-like adipocytes by increasing the expression of white adipocyte-specific genes and repressing the expression of brown adipocyte-specific genes. nlrp3 deletion in pink1 knockout mice reversed BAT dysfunction. Conversely, adipose tissue-specific atg7 KO mice showed significantly lower expression of Nlrp3 in their BAT. Overall, our data suggest that the role of mitophagy is different from general autophagy in regulating adipose tissue and whole-body energy metabolism. Our results uncovered a new mitochondria-NLRP3 pathway that induces BAT dysfunction. The ability of the nlrp3 knockouts to rescue BAT dysfunction suggests the transcriptional function of NLRP3 as an unexpected, but a quite specific therapeutic target for obesity-related metabolic diseases.Abbreviations: ACTB: actin, beta; BAPs: brown adipocyte precursors; BAT: brown adipose tissue; BMDMs: bone marrow-derived macrophages; CASP1: caspase 1; CEBPA: CCAAT/enhancer binding protein (C/EBP), alpha; ChIP: chromatin immunoprecipitation; EE: energy expenditure; HFD: high-fat diet; IL1B: interleukin 1 beta; ITT: insulin tolerance test; KO: knockout; LPS: lipopolysaccharide; NLRP3: NLR family, pyrin domain containing 3; PINK1: PTEN induced putative kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; RD: regular diet; ROS: reactive oxygen species; RT: room temperature; UCP1: uncoupling protein 1 (mitochondrial, proton carrier); WT: wild-type.
UBE4B, a microRNA-9 target gene, promotes autophagy-mediated Tau degradation
No full textThe formation of hyperphosphorylated intracellular Tau tangles in the brain is a hallmark of Alzheimer’s disease (AD). Tau hyperphosphorylation destabilizes microtubules, promoting neurodegeneration in AD patients. To identify suppressors of tau-mediated AD, we perform a screen using a microRNA (miR) library in Drosophila and identify the miR-9 family as suppressors of human tau overexpression phenotypes. CG11070, a miR-9a target gene, and its mammalian orthologue UBE4B, an E3/E4 ubiquitin ligase, alleviate eye neurodegeneration, synaptic bouton defects, and crawling phenotypes in Drosophila human tau overexpression models. Total and phosphorylated Tau levels also decrease upon CG11070 or UBE4B overexpression. In mammalian neuroblastoma cells, overexpression of UBE4B and STUB1, which encodes the E3 ligase CHIP, increases the ubiquitination and degradation of Tau. In the Tau-BiFC mouse model, UBE4B and STUB1 overexpression also increase oligomeric Tau degradation. Inhibitor assays of the autophagy and proteasome systems reveal that the autophagy-lysosome system is the major pathway for Tau degradation in this context. These results demonstrate that UBE4B, a miR-9 target gene, promotes autophagy-mediated Tau degradation together with STUB1, and is thus an innovative therapeutic approach for AD.
Long-term depletion of Cereblon induces mitochondrial dysfunction in cancer cells
No full textCereblon (CRBN) is a multi-functional protein that acts as a substrate receptor of the E3 ligase complex and a molecular chaperone. While CRBN is proposed to function in mitochondria, its specific roles are yet to be established. Here, we showed that knockdown of CRBN triggers oxidative stress and calcium overload in mitochondria, leading to disruption of mitochondrial membrane potential. Notably, long-term CRBN depletion using PROteolysis TArgeting Chimera (PROTAC) induced irreversible mitochondrial dysfunction, resulting in cell death. Our collective findings indicate that CRBN is required for mitochondrial homeostasis in cells.
Anti-inflammatory effect of Barringtonia angusta methanol extract is mediated by targeting of Src in the NF-κB signalling pathway
No full textContext: Among the plants in the genus Barringtonia (Lecythidaceae) used as traditional medicines to treat arthralgia, chest pain, and haemorrhoids in Indonesia, Barringtonia racemosa L. and Barringtonia acutangula (L.) Gaertn. have demonstrated anti-inflammatory activity in systemic inflammatory models.
Objective: The anti-inflammatory activity of Barringtonia angusta Kurz has not been investigated. We prepared a methanol extract of the leaves and stems of B. angusta (Ba-ME) and systemically evaluated its anti-inflammatory effects in vitro and in vivo.
Materials and methods: RAW264.7 cells stimulated with LPS or Pam3CSK4 for 24？h were treated with Ba-ME (12.5, 25, 50, 100, and 150？μg/mL), and NO production and mRNA levels of inflammatory genes were evaluated. Luciferase reporter gene assay, western blot analysis, overexpression experiments, and cellular thermal shift assay were conducted to explore the mechanism of Ba-ME. In addition, the anti-gastritis activity of Ba-ME (50 and 100？mg/kg, administered twice per day for two days) was evaluated using an HCl/EtOH-induced gastritis mouse model.
Results: Ba-ME dose-dependently suppressed NO production [IC50 = 123.33？μg/mL (LPS) and 46.89？μg/mL (Pam3CSK4)] without affecting cell viability. Transcriptional expression of iNOS, IL-1β, COX-2, IL-6, and TNF-α and phosphorylation of Src, IκBα, p50/105, and p65 were inhibited by Ba-ME. The extract specifically targeted the Src protein by binding to its SH2 domain. Moreover, Ba-ME significantly ameliorated inflammatory lesions in the HCl/EtOH-induced gastritis model.
Discussion and Conclusions: The anti-inflammatory activity of Ba-ME is mediated by targeting of the Src/NF-κB signalling pathway, and B. angusta has potential as an anti-inflammatory drug.
Escherichia coli shiga toxins and gut microbiota interactions
No full textEscherichia coli (EHEC) and Shigella dysenteriae serotype 1 are enterohemorrhagic bacteria that induce hemorrhagic colitis. This, in turn, may result in potentially lethal complications, such as hemolytic uremic syndrome (HUS), which is characterized by thrombocytopenia, acute renal failure, and neurological abnormalities. Both species of bacteria produce Shiga toxins (Stxs), a phage-encoded exotoxin inhibiting protein synthesis in host cells that are primarily responsible for bacterial virulence. Although most studies have focused on the pathogenic roles of Stxs as harmful substances capable of inducing cell death and as proinflammatory factors that sensitize the host target organs to damage, less is known about the interface between the commensalism of bacterial communities and the pathogenicity of the toxins. The gut contains more species of bacteria than any other organ, providing pathogenic bacteria that colonize the gut with a greater number of opportunities to encounter other bacterial species. Notably, the presence in the intestines of pathogenic EHEC producing Stxs associated with severe illness may have compounding effects on the diversity of the indigenous bacteria and bacterial communities in the gut. The present review focuses on studies describing the roles of Stxs in the complex interactions between pathogenic Shiga toxin-producing E. coli, the resident microbiome, and host tissues. The determination of these interactions may provide insights into the unresolved issues regarding these pathogens.
Production and application of new Immune control substances using marine microbial immunization system, CRISPR/Cas
No full text해양 미생물 면역체계 CRISPR/Cas 시스템을 이용한 신규 면역 조절 물질 생산과 활용MBC006201
Acetylcholinesterase and monoamine oxidase-B inhibitory activities by ellagic acid derivatives isolated from Castanopsis cuspidata var. sieboldii
No full textAmong 276 herbal extracts, a methanol extract of Castanopsis cuspidata var. sieboldii stems was selected as an experimental source for novel acetylcholinesterase (AChE) inhibitors. Five compounds were isolated from the extract by activity-guided screening, and their inhibitory activities against butyrylcholinesterase (BChE), monoamine oxidases (MAOs), and β-site amyloid precursor protein cleaving enzyme 1 (BACE-1) were also evaluated. Of these compounds, 4'-O-(α-L-rhamnopyranosyl)-3,3',4-tri-O-methylellagic acid (3) and 3,3',4-tri-O-methylellagic acid (4) effectively inhibited AChE with IC50 values of 10.1 and 10.7 μM, respectively. Ellagic acid (5) inhibited AChE (IC50 = 41.7 μM) less than 3 and 4. In addition, 3 effectively inhibited MAO-B (IC50 = 7.27 μM) followed by 5 (IC50 = 9.21 μM). All five compounds weakly inhibited BChE and BACE-1. Compounds 3, 4, and 5 reversibly and competitively inhibited AChE, and were slightly or non-toxic to MDCK cells. The binding energies of 3 and 4 (- 8.5 and - 9.2 kcal/mol, respectively) for AChE were greater than that of 5 (- 8.3 kcal/mol), and 3 and 4 formed a hydrogen bond with Tyr124 in AChE. These results suggest 3 is a dual-targeting inhibitor of AChE and MAO-B, and that these compounds should be viewed as potential therapeutics for the treatment of Alzheimer's disease.