Analysis on the Molecular Biologic Characteristics and Expression of Lysozyme C Separated from Oplegnathus fasciatus

Background and Aim: lysozyme has been known as a significant component of the innate immune system of fish. It is reported that the g-type and c-type lysozymes have been identified in fish. Oplegnathus fasciatus is one of the economically important cultured species, generating higher market value and demands in Korea. Contrary to the heavy consumption of O. fasciatus, however, there has been very limited information on their immune system and immunity genes responding against diseases.Methods: This study clarified the molecular biological characteristics of cDNA of the C-type lysozyme from O. fasicatus and conducted phylogency analysis. Furthermore, tissue-specific expression analysis of genes verified the roles of c-type lysozyme in general circumstances.Results: The full-length RbLysC cDNA was 985 bp long and contained an ORF of 432 bp that encoded 143 aa residues, the 5’-UTR of 120 bp and the 3’-UTR of 444 bp. Signal peptide was not found in the terminal and flanking active aspartate region are conserved in RbLysC, and a polyadenylation signal and poly A-tail are present in the 3’-UTR of RbLysC. RbLysC presented the closest distant relationship with sequences from Yellow perch lysozyme C. The highest RbLysC gene expression was observed in the liver, which was about 50-fold relative to that of the PBLs.</p

Prognosis and Sensitivity of Adjuvant Chemotherapy in Mucinous Colorectal Adenocarcinoma without Distant Metastasis

In colorectal cancer, whereas mucinous adenocarcinoma (MAC) has several poor clinical prognostic factors compared to adenocarcinoma (AC), the prognosis of MAC remains controversial. We evaluated the prognosis of MAC without distant metastasis and the effects of adjuvant chemotherapy using health insurance registry data managed by South Korea. Patients with colorectal cancer between January 2014 and December 2016 were included (AC, 22,050 [96.8%]; MAC, 729 [3.2%]). We observed no difference in overall survival (OS) between AC and MAC in stages I and II. However, MAC showed a worse OS than AC in stage III disease, especially in patients administered chemotherapy (p < 0.001). These findings persisted after propensity score matching of clinical characteristics between AC and MAC. In addition, transcriptome analysis of The Cancer Genome Atlas (TCGA) data showed increased chemoresistance-associated pathways in MAC compared to AC. In consensus molecular subtypes (CMS) classification, unlike in AC, CMSs 1, 3, and 4 comprised most of MAC and the proportions of CMSs 3 and 4 increased with stage progression. These results suggest clues to overcome resistance to chemotherapy and develop targeted treatments in MAC

Role of Hepcidins from Black Rockfish (Sebastes schlegelii) in Iron-Metabolic Function and Bacterial Defense

Hepcidin, an antimicrobial peptide produced by the liver, also controls the iron balance and regeneration in vertebrates. Two types of hepcidin (Hamp1 and Hamp2) have been found in the bodies of black rockfish (Sebastes schlegelii). The full-length cDNA of hepcidin was cloned to enable a study of the antibacterial roles of these two hepcidins (Hamp) in black rockfish. The antimicrobial function of recombinant hepcidins was tested both in vitro and in vivo by the synthesis in Escherichia coli of recombinant hepcidin (approximately 11 kDa) from black rockfish. The recombinant hepcidins inhibited the growth of two bacterial species, Streptococcus iniae FP5228 and Pseudomonas aeruginosa, at various concentrations, in vitro after 6 h post-incubation, respectively. During infection, the production of ferroportin was reduced, suggesting the preservation of iron to prevent microbial proliferation. In vivo administration of Hamp1, but not Hamp2, synthetic peptides induced a substantial reduction in the expression of ferroportin, suggesting that in black rockfish with two forms of hepcidin, ferroportin production is regulated by the iron-regulator Hamp1, and not by the dedicated antimicrobial Hamp2. The findings of this study suggest the various antimicrobial roles of these two types of hepcidin

Characterization, Expression Profile, and Promoter Analysis of the Rhodeus uyekii Vitellogenin Ao1 Gene

The fish Vitellogenin (Vg) gene has been applied as a biomarker for exposure to estrogenic compounds in the aquatic environment. In this study, we cloned and characterized Vg cDNA from the Korean rose bitterling Rhodeus uyekii (Ru-Vg). The Ru-Vg cDNA encodes a 1424-amino-acid polypeptide that belongs to the VgAo1 family and contains a putative signal peptide, lipovitellin I, phosvitin, and lipovitellin II, but does not contain the vWFD domain or the C-terminal peptide. The deduced Ru-Vg protein has high amino acid identity (73.97%–32.17%) with fish Vg proteins. Pairwise alignment and phylogenetic analysis revealed that Ru-Vg is most closely related to Acheilognathus yamatsutae Vg. Ru-Vg transcripts were detected using quantitative polymerase chain reaction in all tissues tested, with the highest level of expression observed in the ovary. Ru-Vg mRNA was upregulated in R. uyekii hepatopancreas cells in response to treatment with 17β-estradiol (E2) or 17α-ethinylestradiol (EE2). Luciferase reporter expression, driven by the 5\u27-regulatory region of the Ru-Vg gene spanning from −1020 bp to the start codon was induced by the estrogen receptor and was synergistically activated by treatment with E2 or EE2. These results suggest that R. uyekii and the Ru-Vg gene may be useful as biomarkers for exposure to E2 or EE2

Molecular and Functional Characterization of Thioredoxin 1from Korean Rose Bitterling (Rhodeus uyekii)

Thioredoxin is a multifunctional antioxidant enzyme that belongs to the reductase family. In this study, we cloned and characterized thioredoxin 1 cDNA from the Korean rose bitterling Rhodeus uyekii (RuTrx). The full-length RuTrx cDNA consists of 674 bp with a 324 nt open reading frame (ORF) encoding a 107 aa protein. The deduced RuTrx amino acid sequence indicated a characteristic redox active site, 31WCGPC35. Pairwise alignment revealed RuTrx amino acid identity (55.1%–83.2%) with orthologs from various species of mammalia, amphibia, fish and bird. Phylogenetic analysis was conducted to determine the evolutionary position of RuTrx. Expression analysis showed that RuTrx transcripts were present in all of the tissues examined, and was high in the hepatopancreas of R. uyekii. During early development, the expression of RuTrx transcripts was increased. Recombinant RuTrx protein (rRuTrx) was tested for its capacity to serve as an antioxidant enzyme using a metal-catalyzed oxidation (MCO) system. The ability of rRuTrx to protect against supercoiled DNA cleavage due to oxidative nicking increased in a dose-dependent manner. In Raw264.7 cells, Dihydroethidium (DHE) staining for ROS production indicated the antioxidant activity of rRuTrx. Together, these findings suggest that RuTrx may play a role in maintaining the redox state balance in Korean rose bitterling R. uyekii

Liquid Chromatography-Mass Spectrometry-Based Rapid Secondary-Metabolite Profiling of Marine Pseudoalteromonas sp. M2

The ocean is a rich resource of flora, fauna, and food. A wild-type bacterial strain showing confluent growth on marine agar with antibacterial activity was isolated from marine water, identified using 16S rDNA sequence analysis as Pseudoalteromonas sp., and designated as strain M2. This strain was found to produce various secondary metabolites including quinolone alkaloids. Using high-resolution mass spectrometry (MS) and nuclear magnetic resonance (NMR) analysis, we identified nine secondary metabolites of 4-hydroxy-2-alkylquinoline (pseudane-III, IV, V, VI, VII, VIII, IX, X, and XI). Additionally, this strain produced two novel, closely related compounds, 2-isopentylqunoline-4-one and 2-(2,3-dimetylbutyl)qunoline-4-(1H)-one, which have not been previously reported from marine bacteria. From the metabolites produced by Pseudoalteromonas sp. M2, 2-(2,3-dimethylbutyl)quinolin-4-one, pseudane-VI, and pseudane-VII inhibited melanin synthesis in Melan-A cells by 23.0%, 28.2%, and 42.7%, respectively, wherein pseudane-VII showed the highest inhibition at 8 µg/mL. The results of this study suggest that liquid chromatography (LC)-MS/MS-based metabolite screening effectively improves the efficiency of novel metabolite discovery. Additionally, these compounds are promising candidates for further bioactivity development

Layer-Confined Excitonic Insulating Phase in Ultrathin Ta2NiSe5 Crystals

Atomically thin nanosheets, as recently realized using van der Waals layered materials, offer a versatile platform for studying the stability and tunability of the correlated electron phases in the reduced dimension. Here, we investigate a thickness-dependent excitonic insulating (EI) phase on a layered ternary chalcogenide Ta2NiSe5. Using Raman spectroscopy, scanning tunneling spectroscopy, and in-plane transport measurements, we found no significant changes in crystalline and electronic structures as well as disorder strength in ultrathin Ta2NiSe5 crystals with a thickness down to five layers. The transition temperature, Tc, of ultrathin Ta2NiSe5 is reduced from its bulk value by ΔTc/Tc bulk ≈ -9%, which strongly contrasts the case of 1T-TiSe2, another excitonic insulator candidate, showing an increase of Tc by ΔTc/Tc bulk ≈ +30%. This difference is attributed to the dominance of interband Coulomb interaction over electron-phonon interaction and its zero-ordering wave vector due to the direct band gap structure of Ta2NiSe5. The out-of-plane correlating length of the EI phase is estimated to have monolayer thickness, suggesting that the EI phase in Ta2NiSe5 is highly layer-confined and in the strong coupling limit. © 2016 American Chemical Society9

Molecular and functional characterizations of a Kunitz-type serine protease inhibitor FcKuSPI of the shrimp Fenneropenaeus chinensis

Serine proteinase inhibitors play important and diverse roles in biological processes such as coagulation, defense mechanisms, and immune responses. Here, we identified and characterized a Kunitz-type proteinase inhibitor, designated FcKuSPI, of the BPTI/Kunitz family of serine proteinase inhibitors from the hemocyte cDNA library of the shrimp Fenneropenaeus chinensis. The deduced amino acid sequence of FcKuSPI comprises 80 residues with a putative signal peptide of 15 amino acids. The predicted molecular weight of the mature peptide is 7.66 kDa and its predicted isoelectric point is 8.84. FcKuSPI includes a Kunitz domain containing six conserved cysteine residues that are predicted to form three disulfide bonds. FcKuSPI shares 44e53% homology with BPTI/Kunitz family members from other species. FcKuSPI mRNAwas expressed highly in the hemocytes and moderately in muscle in healthy shrimp. Recombinant FcKuSPI protein demonstrated anti-protease activity against trypsin and anticoagulant activity against citrated human plasma in a dose-dependent manner in in vitro assays

Transcriptome Analysis Revealed Changes of Multiple Genes Involved in Haliotis discus hannai Innate Immunity during Vibrio parahemolyticus Infection.

Abalone (Haliotis discus hannai) is one of the most valuable marine aquatic species in Korea, Japan and China. Tremendous exposure to bacterial infection is common in aquaculture environment, especially by Vibrio sp. infections. It's therefore necessary and urgent to understand the mechanism of H. discus hannai host defense against Vibrio parahemolyticus infection. However studies on its immune system are hindered by the lack of genomic resources. In the present study, we sequenced the transcriptome of control and bacterial challenged H. discus hannai tissues. Totally, 138 MB of reference transcriptome were obtained from de novo assembly of 34 GB clean bases from ten different libraries and annotated with the biological terms (GO and KEGG). A total of 10,575 transcripts exhibiting the differentially expression at least one pair of comparison and the functional annotations highlight genes related to immune response, cell adhesion, immune regulators, redox molecules and mitochondrial coding genes. Mostly, these groups of genes were dominated in hemocytes compared to other tissues. This work is a prerequisite for the identification of those physiological traits controlling H. discus hannai ability to survive against Vibrio infection