Immunostimulating Effects of Extract of Acanthopanax sessiliflorus

As malfunction/absence of immune cells causes a variety of immunosuppressive disorders and chemical synthetic drugs for curing these diseases have many adverse effects, vigorous studies are being conducted. The Acanthopanax family has been used as traditional medicines for gastric ulcer, diabetes, etc. and culinary materials in East-South Asia. In this study, the immunostimulating properties of A. sessiliflorus were evaluated. A. sessiliflorus increased not only the splenocyte number but also immune-related cytokines such as TNF-α. However, it could not upregulate the expressions of IFN-γ and IL-2. A. sessiliflorus increased the swimming time, and comparison of organ weights relative to body weights for immune-related organs such as the spleen and thymus after a forced swim test showed that it could recover the spleen and thymus weights. It also increased the expression of TNF-α and slightly increased the concentration of IFN-γ but not IL-2. From the results, we concluded that as A. sessiliflorus has not only a host defense effect but also a stress-ameliorating property, further study it will be a promising material of immunostimulating material.     

Biodegradation of endocrine-disrupting bisphenol A by white rot fungus Irpex lacteus

Biodegradation of endocrine-disrupting bisphenol A was investigated with several white rot fungi (Irpex lacteus, Trametes versicolor, Ganoderma lucidum, Polyporellus brumalis, Pleurotus eryngii, Schizophyllum commune) isolated in Korea and two transformants of T versicolor (strains MrP 1 and MrP 13). I. lacteus degraded 99.4% of 50 mg/l bisphenol A in 3 h incubation and 100% in 12 h incubation. which was the highest degradation rate among the fungal strains tested. T. versicolor degraded 98.2% of 50 mg/l bisphenol A in 12 h incubation. Unexpectedly, the transformant of the Mn-repressed peroxidase gene of T. versicolor, strain MrP 1, degraded 76.5% of 50 mg/l bisphenol A in 12 h incubation, which was a lower degradation rate than wild-type T. versicolor. The removal of bisphenol A by I. lacteus occurred mainly by biodegradation rather than adsorption. Optimum carbon sources for biodegradation of bisphenol A by I. lacteus were glucose and starch, and optimum nitrogen sources were yeast extract and tryptone in a minimal salts medium; however, bisphenol A degradation was higher in nutrient-rich YMG medium than that in a minimal salts medium. The initial degradation of endocrine disruptors was accompanied by the activities of manganese peroxidase and laccase in the culture     

The role of p38 MAP kinase and c-Jun N-terminal protein kinase signaling in the differentiation and apoptosis of immortalized neural stem cells

The two distinct members of the mitogen-activated protein (MAP) kinase family c-Jun N-terminal protein kinase (JNK) and p38 MAP kinase, play an important role in central nervous system (CNS) development and differentiation. However, their role and functions are not completely understood in CNS. To facilitate in vitro study, we have established an immortal stem cell line using SV40 from fetal rat embryonic day 17. In these cells, MAP kinase inhibitors (SP600125, SB202190, and PD98059) were treated for 1, 24, 48, and 72 h to examine the roles of protein kinases. Early inhibition of JNK did not alter phenotypic or morphological changes of immortalized cells, however overexpression of Bax and decrease of phosphorylated AKT was observed. The prolonged inhibition of JNK induced polyploidization of immortalized cells, and resulted in differentiation and inhibition of cell proliferation. Moreover, JNK and p38 MAP kinase but not ERK1/2 was activated, and p21, p53, and Bax were overexpressed by prolonged inhibition of JNK.

These results indicate that JNK and p38 MAP kinase could play dual roles on cell survival and apoptosis. Furthermore, this established cell line could facilitate study of the role of JNK and p38 MAP kinase on CNS development or differentiation/apoptosis.     

Molecular phylogeny and geography of Korean medaka fish (Oryzias latipes)

The phylogeny and geography of the medaka (Oryzias latipes) populations of Korea were investigated by analyzing sequence data for the mitochondrial control region. From the 41 haplotypes including 25 Korean haplotypes detected in 64 Korean specimens and data for the Japanese and Chinese populations, phylogenetic and nested clade analyses were executed to examine the phylogeny of haplogroups and the relation of the genetic architecture of the haplotypes to the historical geography of the Korean medaka fish. The analyses suggest that there are two very distinct lineages of Korean medaka, and that these result from reproductive isolation mechanisms due to geographic barriers. The southeastern lineage has experienced recent range expansion to the western region. The northwestern lineage, sister to Chinese populations, showed evidence of internal range expansion with shared haplotypes.     

Snx5, as a Mind bomb-binding protein, is expressed in hematopoietic and endothelial precursor cells in zebrafish

Notch signaling has an evolutionarily conserved function for cell fate determination and stem cell maintenance. Previously, we identified a novel component of the Notch signaling pathway in zebrafish, mind bomb, which encodes an E3 ubiquitin ligase essential for Notch signal activation. Further studies showed that Mind bomb(-/-) mouse embryos exhibited pan-Notch phenotypes in various tissues, suggesting that Mind bomb function is conserved in mammals. Therefore we sought to understand the various molecular partners of Mind bomb using yeast two-hybrid screening. In this search we identified Sorting nexin 5 (Snx5) as a novel interacting partner of Mind bomb. Furthermore we demonstrated that Snx5 colocalizes with Mind bomb in early endosomal compartments, suggesting that Snx5 is important for Mind bomb trafficking. In addition, we identified zebrafish orthologue of Snx5 and showed that snx5 is predominantly expressed in hematopoietic and endothelial precursor cells in zebrafish. We also found defects in hematopoiesis and blood vessel development in snx5 morpholino-injected embryos. Taken together, we show that Snx5, a novel interacting partner of Mind bomb, may have an essential role for cell fate determination in early development.     

93-kDa twin-domain serine protease inhibitor (Serpin) has a regulatory function on the beetle Toll proteolytic signaling cascade

Serpins are protease inhibitors that play essential roles in the down-regulation of extracellular proteolytic cascades. The core serpin domain is highly conserved, and typical serpins are encoded with a molecular size of 35–50 kDa. Here, we describe a novel 93-kDa protein that contains two complete, tandemly arrayed serpin domains. This twin serpin, SPN93, was isolated from the larval hemolymph of the large beetle Tenebrio molitor. The N-terminal serpin domain of SPN93 forms a covalent complex with the Spätzle-processing enzyme, a terminal serine protease of the Toll signaling cascade, whereas the C-terminal serpin domain of SPN93 forms complexes with a modular serine protease and the Spätzle-processing enzyme-activating enzyme, which are two different enzymes of the cascade. Consequently, SPN93 inhibited β-1,3-glucan-mediated Toll proteolytic cascade activation in an in vitro system. Site-specific proteolysis of SPN93 at the N-terminal serpin domain was observed after activation of the Toll proteolytic cascade in vivo, and down-regulation of SPN93 by RNAi sensitized β-1,3-glucan-mediated larval death. Therefore, SPN93 is the first serpin that contains twin tandemly arrayed and functionally active serpin domains that have a regulatory role in the larval Toll proteolytic signaling cascade.     

Structural and functional characterization of a highly specific serpin in the insect innate immunity

The Toll signaling pathway, an essential innate immune response in invertebrates, is mediated via the serine protease cascade. Once activated, the serine proteases are irreversibly inactivated by serine protease inhibitors (serpins). Recently, we identified three serpin-serine protease pairs that are directly involved in the regulation of Toll signaling cascade in a large beetle, Tenebrio molitor. Of these, the serpin SPN48 was cleaved by its target serine protease, Spätzle-processing enzyme, at a noncanonical P1 residue of the serpin''s reactive center loop. To address this unique cleavage, we report the crystal structure of SPN48, revealing that SPN48 exhibits a native conformation of human antithrombin, where the reactive center loop is partially inserted into the center of the largest β-sheet of SPN48. The crystal structure also shows that SPN48 has a putative heparin-binding site that is distinct from those of the mammalian serpins. Ensuing biochemical studies demonstrate that heparin accelerates the inhibition of Spätzle-processing enzyme by a proximity effect in targeting the SPN48. Our finding provides the molecular mechanism of how serpins tightly regulate innate immune responses in invertebrates.