http://chineseinput.net/에서 pinyin(병음)방식으로 중국어를 변환할 수 있습니다.
변환된 중국어를 복사하여 사용하시면 됩니다.
Sandamalika, W.M. Gayashani,Priyathilaka, Thanthrige Thiunuwan,Nam, Bo-Hye,Lee, Jehee ACADEMIC PRESS LTD 2019 FISH AND SHELLFISH IMMUNOLOGY Vol.87 No.-
<P><B>Abstract</B></P> <P>Phospholipid scramblases (PLSCRs) are a family of transmembrane proteins known to be responsible for Ca<SUP>2+</SUP>-mediated bidirectional phospholipid translocation in the plasma membrane. Apart from the scrambling activity of PLSCRs, recent studies revealed their diverse other roles, including antiviral defense, tumorigenesis, protein–DNA interactions, apoptosis regulation, and cell activation. Nonetheless, the biological and transcriptional functions of PLSCRs in fish have not been discovered to date. Therefore, in this study, two new members related to the PLSCR1 family were identified in the red lip mullet (<I>Liza haematocheila</I>) as <I>MuPLSCR1like-a</I> and <I>MuPLSCR1like-b</I>, and their characteristics were studied at molecular and transcriptional levels. Sequence analysis revealed that MuPLSCR1like-a and MuPLSCR1like-b are composed of 245 and 228 amino acid residues (aa) with the predicted molecular weights of 27.82 and 25.74 kDa, respectively. A constructed phylogenetic tree showed that MuPLSCR1like-a and MuPLSCR1like-b are clustered together with other known PLSCR1 and -2 orthologues, thus pointing to the relatedness to both PLSCR1 and PLSCR2 families. Two-dimensional (2D) and 3D graphical representations illustrated the well-known 12-stranded β-barrel structure of MuPLSCR1like-a and MuPLSCR1like-b with transmembrane orientation toward the phospholipid bilayer. In analysis of tissue-specific expression, the highest expression of <I>MuPLSCR1like-a</I> was observed in the intestine, whereas <I>MuPLSCR1like-b</I> was highly expressed in the brain, indicating isoform specificity. Of note, we found that the transcription of <I>MuPLSCR1like-a</I> and <I>MuPLSCR1like-b</I> was significantly upregulated when the fish were stimulated with poly(I:C), suggesting that such immune responses target viral infections. Overall, this study provides the first experimental insight into the characteristics and immune-system relevance of <I>PLSCR1</I>-related genes in red lip mullets.</P> <P><B>Highlights</B></P> <P> <UL> <LI> <I>MuPLSCR1like-a</I> and <I>MuPLSCR1like-b</I> were identified from red lip mullet. </LI> <LI> Highest expression of <I>MuPLSCR1like-a</I> was observed in intestine. </LI> <LI> <I>MuPLSCR1like-b</I> was highly expressed in brain. </LI> <LI> Both genes were significantly upregulated after stimulated with poly(I:C). </LI> </UL> </P>
Sandamalika, W.M. Gayashani,Priyathilaka, Thanthrige Thiunuwan,Lee, Seongdo,Yang, Hyerim,Lee, Jehee ACADEMIC PRESS LTD 2019 FISH AND SHELLFISH IMMUNOLOGY Vol.91 No.-
<P><B>Abstract</B></P> <P>Representing a multifunctional complex group of proteins, glutathione S- transferases (GSTs) play a major role in the phase II detoxification process in a wide range of organisms. This study focused on the potential detoxification ability of disk abalone (<I>Haliotis discus discus</I>) GST theta (<I>AbGST-θ</I>) under different stress conditions with special reference to post immune challenges. Characterization of AbGST-θ revealed with 226 amino acids, 26.6 kDa of predicted molecular mass and 8.9 of theoretical isoelectric point. As illustrated in the multiple sequence alignment, eight glutathione binding sites (G-sites) and ten substrate binding sites (H-sites) were identified in well-distinct N-terminal and C-terminal domains of AbGST-θ, respectively. AbGST-θ exhibited its highest sequence identity with <I>Mizuhopecten yessoensis</I> (59.1%) and the phylogenetic tree clearly positioned AbGST-θ with pre-defined GST-θ molluscan homologues. The <I>AbGST-θ</I> was highly expressed in the digestive tract of un-challenged abalones. Upon administering the challenge experiment, <I>AbGST-θ</I> showed significant modulations in their transcriptional levels depending on the time and the tissue type. The optimum temperature was 37 °C and optimum pH was 7.5 for AbGST-θ. The determined enzyme kinetic parameters of AbGST-θ showed low affinity towards 1-Chloro-2,4-dinitrobenzene (CDNB) and glutathione (GSH) as substrates. Nonetheless, with Cibacron blue IC<SUB>50</SUB> (half maximal inhibitory concentration) was calculated to be 0.08 μM while observing 100% inhibition with 100 μM. Furthermore, AbGST-θ resulted in significant protection ability towards H<SUB>2</SUB>O<SUB>2</SUB>, CdCl<SUB>2,</SUB> and ZnCl<SUB>2</SUB> in the disk diffusion assay. Collectively, this study provides evidences for the detoxification ability and the immunological host defensive capability of <I>AbGST-θ</I> in disk abalone<I>.</I> </P> <P><B>Highlights</B></P> <P> <UL> <LI> We identified Glutathione S-transferases theta from disk abalone (AbGST-θ). </LI> <LI> AbGST-θ revealed with 226 amino acids. </LI> <LI> <I>AbGST-θ</I> was highly expressed in the digestive tract of un-challenged abalones. </LI> <LI> <I>AbGST-θ</I> mRNA expression level significantly increased after immune challenge. </LI> <LI> Enzyme kinetics, optimum conditions and heavy metal stress responses were measured. </LI> </UL> </P>
Sandamalika, W.M. Gayashani,Priyathilaka, Thanthrige Thiunuwan,Liyanage, D.S.,Lee, Sukkyoung,Lim, Han-Kyu,Lee, Jehee Elsevier 2018 FISH AND SHELLFISH IMMUNOLOGY Vol.77 No.-
<P><B>Abstract</B></P> <P>Glutathione S-transferase (GST; EC 2.5.1.18) isoenzymes represent a complex group of proteins that are involved in phase II detoxification in several organisms. In this study, GST kappa (GSTκ) from the disk abalone (<I>Haliotis discus discus; AbGSTκ</I>) was characterized at both the transcriptional and functional levels to determine its potential capacity to perform as a detoxification agent under conditions of different stress. The predicted AbGSTκ protein consists of 227 amino acids, with a predicted molecular weight of 25.6 kDa and a theoretical isoelectric point (pI) of 7.78. <I>In silico</I> analysis reveals that <I>AbGSTκ</I> is a disulfide bond formation protein A (DsbA), consisting of a thioredoxin domain, GSH binding sites (G-sites), and a catalytic residue. In contrast, no hydrophobic ligand binding site (H-site), or signal peptides, were detected. <I>AbGSTκ</I> showed the highest sequence identity with the orthologue from pufferfish (<I>Takifugu obscurus</I>) (60.0%). In a phylogenetic tree, AbGSTκ clustered closely together with other fish GSTκs, and was evolutionarily distanced from other cytosolic GSTs. The predicted three-dimensional structure clearly demonstrates that the dimer adopts a butterfly-like shape. A tissue distribution analysis revealed that <I>GSTκ</I> was highly expressed in the digestive tract, suggesting it has detoxification ability. Depending on the tissue and time, <I>AbGSTκ</I> showed different expression patterns, and levels of expression, following challenge of the abalone with immune stimulants. Enzyme kinetics of the purified recombinant proteins demonstrated its conjugating ability using 1-Chloro-2,4-dinitrobenzene (CDNB) and glutathione (GSH) as substrates, and suggested it has a low affinity for both substrates. The optimum temperature and pH for the rAbGSTκ GSH: CDNB conjugating activity were found to be 35 °C and pH 8, respectively indicating that the abalone is well adapted to a wide range of environmental conditions. Cibacron blue (100 μM) was capable of completely inhibiting rAbGSTκ (100%) with an IC<SUB>50</SUB> (half maximal inhibitory concentration) of 0.05 μM. A disk diffusion assay revealed that rAbGSTκ could significantly protect cells from H<SUB>2</SUB>O<SUB>2</SUB>, CdCl<SUB>2,</SUB> and ZnCl<SUB>2</SUB>. Altogether, this current study suggests that <I>AbGSTκ</I> is involved in detoxification and immunological host defense mechanisms and allows abalones to overcome stresses in order for them to have an increased chance of survival.</P> <P><B>Highlights</B></P> <P> <UL> <LI> The full-length coding sequence of <I>AbGSTκ</I> was identified in disk abalone. </LI> <LI> <I>AbGSTκ</I> is a disulfide bond formation protein A (DsbA) with a thioredoxin domain. </LI> <LI> Highest mRNA expression of AbGSTκ was observed in digestive tract. </LI> <LI> Immunological responses of AbGSTκ has evaluated. </LI> <LI> Enzyme kinetics, optimum conditions and heavy metal stress responses were measured. </LI> </UL> </P>
W.M. Gayashani Sandamalika,Jehee Lee 제주대학교 해양과학연구소 2020 해양과환경연구소 연구논문집 Vol.44 No.-
Galectins are well-known β-galactoside-binding proteins, which play vital roles in innate immune responses of both vertebrates and invertebrates. However, knowledge regarding invertebrate galectins is still in its infancy. With the intention of filling the knowledge gap, here we identified a quadruple domain-containing galectin from marine invertebrate disk abalone, Haliotis discus discus (AbGalec), and characterized it. AbGalec consisted of four distinct carbohydrate-recognition domains (CRDs) and lacked a signal peptide. Expression analysis revealed AbGalec to be ubiquitously expressed in all the examined early embryonic stages of abalone, with highest expression in the 16-cell stage, suggesting the importance of AbGalec in early developmental processes. Tissue distribution analysis revealed the highest expression of AbGalec in abalone mantle, followed by that in gills and hemocytes. Immune challenge experiments revealed significant upregulation of AbGalec at 24 h and 48 h post injection (p.i.) with bacterial and viral components. These results suggested the possible involvement of AbGalec in host defense mechanisms. Polyinosinic: polycytidylic acid (Poly I:C) and viral hemorrhagic septicemia virus (VHSV) injections were capable of inducing AbGalec transcript expression more prominently than bacterial stimulants, thus providing evidence for its role in viral infections. We determined the virus-neutralizing ability of a quadruple domain-containing galectin for the first time, by analyzing the downregulation of VHSV transcripts during the overexpression of AbGalec. Significant downregulation of VHSV transcripts was observed after 24 h and 48 h of post infection. Collectively, our findings reveal the potent antiviral responses of molluscan quadruple domain-containing galectin, AbGalec, along with its involvement in innate immunity.
W.M. Gayashani Sandamalika,Anushka Vidurangi Samaraweera,Hyerim Yang,Jehee Lee 제주대학교 해양과학연구소 2021 해양과환경연구소 연구논문집 Vol.45 No.-
The thioredoxin domain containing 5 (TXNDC5) is a recently discovered member of the protein disulfide isomerase family (PDI), which is mainly involved in the proper folding of and the correct formation of disulfide bonds in newly synthesized proteins via its disulfide isomerase and chaperone activities. Although the structural and functional features of mammalian TXNDC5 have been explored in previous studies, no studies have reported the functional characteristics of TXNDC5 in teleost fish. In this study, we report the identification and characterization of TXNDC5 from big-belly seahorse (Hippocampus abdominalis) (ShTXNDC5) accompanied by functional studies. The in-silico analysis revealed that the gene encodes a 433 amino acid (aa) long polypeptide chain with a predicted molecular weight of 49.3 kDa. According to homology analysis, ShTXNDC5 shares more than 55% sequence similarity with other teleost TXNDC5 proteins, and the alignment of the gene sequence convincingly reflects the accepted phylogeny of teleost. Analysis of the spatial distribution of ShTXNDC5 expression showed that its highest expression was observed in the ovary, gill, and pouch of seahorses. Moreover, significant upregulation of ShTXNDC5 transcription was noted in seahorse blood and kidney tissues in a timedependent manner upon viral and bacterial immune challenges. Furthermore, considerable NADPH turnover, insulin reduction ability and significant cell survival effects of ShTXNDC5 were determined by the functional assay, revealing its capability to overcome cellular oxidative stress. Altogether, these findings expand our understanding of TXNDC5 at the molecular and functional levels, and its putative role in seahorse immunity.