記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Proceedings of the National Academy of Sciences of the United States of America  

DOI： 10.1073/pnas.2121469119

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記述言語：英語   掲載種別：論文集(書籍)内論文   出版者・発行元：Comprehensive Glycoscience: Second Edition  

DOI： 10.1016/B978-0-12-819475-1.00092-4

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.asd.2021.101046

PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1645/19-21

PubMed

記述言語：日本語   出版者・発行元：Frontiers in Chemistry  

Both vertebrates and invertebrates display active innate immune mechanisms for defense against microbial infection, including diversified repertoires of soluble and cell-associated lectins that can effect recognition and binding to potential pathogens, and trigger downstream effector pathways that clear them from the host internal milieu. Galectins are widely distributed and highly conserved lectins that have key regulatory effects on both innate and adaptive immune responses. In addition, galectins can bind to exogenous (“non-self”) carbohydrates on the surface of bacteria, enveloped viruses, parasites, and fungi, and function as recognition receptors and effector factors in innate immunity. Like most invertebrates, eastern oysters (Crassostrea virginica) and softshell clams (Mya arenaria) can effectively respond to most immune challenges through soluble and hemocyte-associated lectins. The protozoan parasite Perkinsus marinus, however, can infect eastern oysters and cause “Dermo” disease, which is highly detrimental to both natural and farmed oyster populations. The sympatric Perkinsus chesapeaki, initially isolated from infected M. arenaria clams, can also be present in oysters, and there is little evidence of pathogenicity in either clams or oysters. In this review, we discuss selected observations from our studies on the mechanisms of Perkinsus recognition that are mediated by galectin-carbohydrate interactions. We identified in the oyster two galectins that we designated CvGal1 and CvGal2, which strongly recognize P. marinus trophozoites. In the clam we also identified galectin sequences, and focused on one (that we named MaGal1) that also recognizes Perkinsus species. Here we describe the biochemical characterization of CvGal1, CvGal2, and MaGal1 with focus on the detailed study of the carbohydrate specificity, and the glycosylated moieties on the surfaces of the oyster hemocytes and the two Perkinsus species (P. marinus and P. chesapeaki). Our goal is to gain further understanding of the biochemical basis for the interactions that lead to recognition and opsonization of the Perkinsus trophozoites by the bivalve hemocytes. These basic studies on the biology of host-parasite interactions may contribute to the development of novel intervention strategies for parasitic diseases of biomedical interest.

DOI： 10.3389/fchem.2020.00098

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記述言語：日本語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Genes  

DOI： 10.3390/genes10121027

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PubMed

担当区分：筆頭著者,　責任著者   記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Molecular Immunology  

DOI： 10.1016/j.molimm.2019.07.020

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PLoS ONE  

There is increasing evidence for frequent turnover in sex chromosomes in vertebrates. Yet experimental systems suitable for tracing the detailed process of turnover are rare. In theory, homologous turnover is possible if the new sex-determining locus is established on the existing sex-chromosome. However, there is no empirical evidence for such an event. The genus Takifugu includes fugu (Takifugu rubripes) and its two closely-related species whose sex is most likely determined by a SNP at the Amhr2 locus. In these species, males are heterozygous, with G and C alleles at the SNP site, while females are homozygous for the C allele. To determine if a shift in the sex-determining locus occurred in another member of this genus, we used genetic mapping to characterize the sex-chromosome systems of Takifugu niphobles. We found that the G allele of Amhr2 is absent in T. niphobles. Nevertheless, our initial mapping suggests a linkage between the phenotypic sex and the chromosome 19, which harbors the Amhr2 locus. Subsequent high-resolution analysis using a sex-reversed fish demonstrated that the sex-determining locus maps to the proximal end of chromosome 19, far from the Amhr2 locus. Thus, it is likely that homologous turnover involving these species has occurred. The data also showed that there is a male-specific reduction of recombination around the sex-determining locus. Nevertheless, no evidence for sex-chromosome differentiation was detected: the reduced recombination depended on phenotypic sex rather than genotypic sex; no X- or Y-specific maker was obtained; the YY individual was viable. Furthermore, fine-scale mapping narrowed down the new sex-determining locus to the interval corresponding to approximately 300-kb of sequence in the fugu genome. Thus, T. niphobles is determined to have a young and small sex-determining region that is suitable for studying an early phase of sex-chromosome evolution and the mechanisms underlying turnover of sex chromosome.

DOI： 10.1371/journal.pone.0190635

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記述言語：日本語   出版者・発行元：JAPANESE SOC FISHERIES SCIENCE  

DOI： 10.2331/suisan.WA2440-1

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Journal of Immunology  

How parasites recognize their definitive hosts is a mystery; however, parasitism is reportedly initiated by recognition of certain molecules on host surfaces. Fish ectoparasites make initial contact with their hosts at body surfaces, such as skin and gills, which are covered with mucosa that are similar to those of mammalian guts. Fish are among the most primitive vertebrates with immune systems that are equivalent to those in mammals, and they produce and secrete IgM into mucus. In this study, we showed that the monogenean parasite Heterobothrium okamotoi utilizes IgM to recognize its host, fugu Takifugu rubripes. Oncomiracidia are infective larvae of H. okamotoi that shed their cilia and metamorphose into juveniles when exposed to purified D-mannose-binding fractions from fugu mucus. Using liquid chromatography-tandem mass spectrometry analysis, proteins contained in the fraction were identified as D-mannose-specific IgM with two D-mannose-binding lectins. However, although deciliation was significantly induced by IgM and was inhibited by D-mannose or a specific Ab against fugu IgM, other lectins had no effect, and IgM without D-mannose affinity induced deciliation to a limited degree. Subsequent immunofluorescent staining experiments showed that fugu D-mannose-specific IgM binds ciliated epidermal cells of oncomiracidium. These observations suggest that deciliation is triggered by binding of fugu IgM to cell surface Ags via Ag binding sites. Moreover, concentrations of D-mannose-binding IgM in gill mucus were sufficient to induce deciliation in vitro, indicating that H. okamotoi parasites initially use host Abs to colonize host gills.

DOI： 10.4049/jimmunol.1601996

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その他リンク： http://orcid.org/0000-0003-3047-6942

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Developmental and Comparative Immunology  

Pufflectin found in Takifugu rubripes (Tr pufflectin) is the first animal lectin reported to show sequence similarity to monocotyledonous plant lectins. In the present study, we identified and characterized an orthologous lectin from Takifugu niphobles (Tn pufflectin), a species closely related to T. rubripes. Tn pufflectin exhibits 86% identity to Tr pufflectin with two conserved mannose-binding domains. Tn pufflectin was mainly expressed in the skin, gills, brain, and muscles; however, it was expressed at a lower level in the other examined tissues. Recombinant Tn pufflectin, expressed by Escherichia coli, exhibited binding activity specific for d-mannose. The expression of pufflectin in the gills was much lower in T. niphobles than in T. rubripes; notably, the former and latter are resistant and susceptible, respectively, to the monogenean parasite Heterobothrium okamotoi, which parasitizes gills. This suggests that pufflectin might be utilized by the parasite for host recognition.

DOI： 10.1016/j.dci.2016.01.007

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記述言語：英語   出版者・発行元：Elsevier {BV}  

DOI： 10.1016/j.fsi.2016.03.134

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Fish and Shellfish Immunology  

Galectins constitute a conserved and widely distributed lectin family characterized by their binding affinity for β-galactosides and a unique binding site sequence motif in the carbohydrate recognition domain (CRD). In spite of their structural conservation, galectins display a remarkable functional diversity, by participating in developmental processes, cell adhesion and motility, regulation of immune homeostasis, and recognition of glycans on the surface of viruses, bacteria and protozoan parasites. In contrast with mammals, and other vertebrate and invertebrate taxa, the identification and characterization of bona fide galectins in aquatic mollusks has been relatively recent. Most of the studies have focused on the identification and domain organization of galectin-like transcripts or proteins in diverse tissues and cell types, including hemocytes, and their expression upon environmental or infectious challenge. Lectins from the eastern oyster Crassostrea virginica, however, have been characterized in their molecular, structural and functional aspects and some notable features have become apparent in the galectin repertoire of aquatic mollusks. These including less diversified galectin repertoires and different domain organizations relative to those observed in vertebrates, carbohydrate specificity for blood group oligosaccharides, and up regulation of galectin expression by infectious challenge, a feature that supports their proposed role(s) in innate immune responses. Although galectins from some aquatic mollusks have been shown to recognize microbial pathogens and parasites and promote their phagocytosis, they can also selectively bind to phytoplankton components, suggesting that they also participate in uptake and intracellular digestion of microalgae. In addition, the experimental evidence suggests that the protozoan parasite Perkinsus marinus has co-evolved with the oyster host to be selectively recognized by the oyster hemocyte galectins over algal food or bacterial pathogens, thereby subverting the oyster's innate immune/feeding recognition mechanisms to gain entry into the host cells.

DOI： 10.1016/j.fsi.2015.05.012

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その他リンク： http://orcid.org/0000-0003-3047-6942

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Biochemistry  

Galectins are highly conserved lectins that are key to multiple biological functions, including pathogen recognition and regulation of immune responses. We previously reported that CvGal1, a galectin expressed in phagocytic cells (hemocytes) of the eastern oyster (Crassostrea virginica), is hijacked by the parasite Perkinsus marinus to enter the host, where it causes systemic infection and death. Screening of an oyster hemocyte cDNA library revealed a novel galectin, which we designated CvGal2, with four tandemly arrayed carbohydrate recognition domains (CRDs). Phylogentic analysis of the CvGal2 CRDs suggests close relationships with homologous CRDs from CvGal1. Glycan array analysis, however, revealed that, unlike CvGal1 which preferentially binds to the blood group A tetrasaccharide, CvGal2 recognizes both blood group A and B tetrasaccharides and related structures, suggesting that CvGal2 has broader binding specificity. Furthermore, SPR analysis demonstrated significant differences in the binding kinetics of CvGal1 and CvGal2, and structural modeling revealed substantial differences in their interactions with the oligosaccharide ligands. CvGal2 is homogeneously distributed in the hemocyte cytoplasm, is released to the extracellular space, and binds to the hemocyte surface. CvGal2 binds to P. marinus trophozoites in a dose-dependent and β-galactoside-specific manner. Strikingly, negligible binding of CvGal2 was observed for Perkinsus chesapeaki, a sympatric parasite species mostly prevalent in the clams Mya arenaria and Macoma balthica. The differential recognition of Perkinsus species by the oyster galectins is consistent with their relative prevalence in oyster and clam species and supports their role in facilitating parasite entry and infectivity in a host-preferential manner.

DOI： 10.1021/acs.biochem.5b00362

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Fish and Shellfish Immunology  

Caligus fugu is a parasitic copepod specific to the tetraodontid genus Takifugu including the commercially important Takifugu rubripes. Despite the rapid accumulation of knowledge on other aspects of its biology, the host and settlement-site recognition mechanisms of this parasite are not yet well understood. Since the infective copepodid stage shows preferential site selection in attaching to the fins, we considered it likely that the copepodid recognizes chemical cues released or leaking from the fins, and/or transmembrane protein present on the fins. To isolate molecules potentially related to attachment site specificity, we applied suppression subtractive hybridization (SSH) PCR by identifying genes expressed more highly in pectoral fins of T.rubripes than in the body surface skin. We sequenced plasmid DNA from 392 clones in a SSH library. The number of non-redundant sequences was 276, which included 135 sequences located on 117 annotated genes and 141 located in positions where no genes had been annotated. We characterized those annotated genes on the basis of gene ontology terms, and found that 46 of the identified genes encode secreted proteins, enzymes or membrane proteins. Among them nine showed higher expression in the pectoral fins than in the skin. These could be candidate genes for involvement in behavioral mechanisms related to the site specificity shown by the infective copepodids of C.fugu.

DOI： 10.1016/j.fsi.2015.02.031

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記述言語：日本語   出版者・発行元：公益社団法人 日本水産学会  

DOI： 10.2331/suisan.81.636

CiNii Research

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Fisheries Science  

The tiger pufferfish (fugu) is one of the most important food fishes in East Asia. Since its testes are regarded as a delicacy, sex determination is economically relevant. Previous studies have identified a missense single-nucleotide polymorphism (SNP) in the Amhr2 (anti-Müllerian hormone receptor type II) gene as a strong candidate for a master sex-determining polymorphism. To distinguish genotypic sex efficiently, we developed a high-resolution melting (HRM) assay for this SNP site. By screening 396 fish from two independent crosses reared under controlled conditions, we observed perfect concordance between the SNP genotype and phenotypic sex. Thus, this method holds great potential for use in high-throughput sexing. When analyzing 293 progeny from a third cross reared under unknown conditions, we unexpectedly found that 25 % of phenotypic males exhibited female genotype. These results suggest that environmental factors such as rearing conditions could influence the sex-determination pathway in pufferfish. Alternatively, genetic modifiers might override the signals from Amhr2. This finding raises a concern regarding enhanced stock management of this species, because sex-reversed fish could compromise the sex ratio in subsequent generations. The HRM assay will also be useful for monitoring the degree of sex reversal before release.

DOI： 10.1007/s12562-014-0768-0

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Parasite  

The complete life cycle of a pennellid copepod Peniculus minuticaudae Shiino, 1956 is proposed based on the discovery of all post-embryonic stages together with the post-metamorphic adult females infecting the fins of threadsail filefish Stephanolepis cirrhifer (Monacanthidae) cultured in a fish farm at Ehime Prefecture, JPN. The hatching stage was the infective copepodid. The life cycle of P. minuticaudae consists of six stages separated by moults: the copepodid, four chalimi and adult. In this study, the adult males were observed frequently in precopulatory amplexus with various stages of females however, copulation occurs only between adults. Fertilized pre-metamorphic adult females carrying spermatophores may detach from the host and settle again before undergoing massive differential growth into the post-metamorphic adult female. Comparison of the life cycle of P. minuticaudae has been made with three known pennellids: Lernaeocera branchialis (Linnaeus, 1767), Cardiodectes medusaeus (Wilson, 1908) and Lernaeenicus sprattae (Sowerby, 1806). Among the compared species, P. minuticaudae is the first ectoparasitic pennellid to be discovered to complete its life cycle on a single host without any change in infection site preferences between infective copepodid and fertilized pre-metamorphic female. © 2013 N. Ismail et al., published by EDP Sciences.

DOI： 10.1051/parasite/2013041

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記述言語：日本語   出版者・発行元：Japanese Society of Animal Breeding and Genetics  

DOI： 10.5924/abgri.41.37

CiNii Research

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PLoS Genetics  

Heterogametic sex chromosomes have evolved independently in various lineages of vertebrates. Such sex chromosome pairs often contain nonrecombining regions, with one of the chromosomes harboring a master sex-determining (SD) gene. It is hypothesized that these sex chromosomes evolved from a pair of autosomes that diverged after acquiring the SD gene. By linkage and association mapping of the SD locus in fugu (Takifugu rubripes), we show that a SNP (C/G) in the anti-Müllerian hormone receptor type II (Amhr2) gene is the only polymorphism associated with phenotypic sex. This SNP changes an amino acid (His/Asp384) in the kinase domain. While females are homozygous (His/His384), males are heterozygous. Sex in fugu is most likely determined by a combination of the two alleles of Amhr2. Consistent with this model, the medaka hotei mutant carrying a substitution in the kinase domain of Amhr2 causes a female phenotype. The association of the Amhr2 SNP with phenotypic sex is conserved in two other species of Takifugu but not in Tetraodon. The fugu SD locus shows no sign of recombination suppression between X and Y chromosomes. Thus, fugu sex chromosomes represent an unusual example of proto-sex chromosomes. Such undifferentiated X-Y chromosomes may be more common in vertebrates than previously thought. © 2012 Kamiya et al.

DOI： 10.1371/journal.pgen.1002798

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記述言語：日本語   出版者・発行元：Springer Nature  

The variable lymphocyte receptors (VLRs) of lamprey and hagfish comprise leucine-rich repeat modules, instead of the immunoglobulin-like domain building blocks of antibodies and T-cell receptors in jawed vertebrates. Both types of vertebrate-rearranging antigen receptors are similarly diverse, with repertoires that can potentially exceed 1014 unique receptors. In order to characterize antigen-binding properties of the VLRs, we developed a high-throughput yeast surface display platform for the isolation of monoclonal VLRs. We have isolated VLRs that specifically bind hen egg lysozyme, β-galactosidase, cholera toxin subunit B, R-phycoerythrin, and the blood group trisaccharides A and B, with binding affinities in the mid-nanomolar to mid-picomolar range. VLRs may, thus, be excellent single-chain alternatives to Ig-based antibodies for biotechnology applications.

DOI： 10.1007/978-1-61779-139-0_2

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Proceedings of the National Academy of Sciences of the United States of America  

Adaptive immunity in jawless vertebrates is mediated by leucine-rich repeat proteins called "variable lymphocyte receptors" (VLRs). Two types of VLR (A and B) are expressed by mutually exclusive lymphocyte populations in lamprey. VLRB lymphocytes resemble the B cells of jawed vertebrates; VLRA lymphocytes are similar to T cells. We determined the structure of a high-affinity VLRA isolated from lamprey immunized with hen egg white lysozyme (HEL) in unbound and antigen-bound forms. The VLRA-HEL complex demonstrates that certain VLRAs, like γδ T-cell receptors (TCRs) but unlike αβ TCRs, can recognize antigens directly, without a requirement for processing orantigen-presenting molecules. Thus, these VLRAs feature the nanomolar affinities of antibodies, the direct recognition of unprocessed antigens of both antibodies and γδ TCRs, and the exclusive expression on the lymphocyte surface that is unique to αβ and γδ TCRs.

DOI： 10.1073/pnas.1005475107

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その他リンク： http://orcid.org/0000-0003-3047-6942

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Proceedings of the National Academy of Sciences of the United States of America  

Lamprey are members of the ancestral vertebrate taxon (jawless fish), which evolved rearranging antigen receptors convergently with the jawed vertebrates. But instead of Ig superfamily domains, lamprey variable lymphocyte receptors (VLRs) consist of highly diverse leucine-rich repeats. Although VLRs represent the only known adaptive immune system not based on Ig, little is known about their antigen-binding properties. Here we report robust plasma VLRB responses of lamprey immunized with hen egg lysozyme and β-galactosidase (β-gal), demonstrating adaptive immune responses against soluble antigens. To isolate monoclonal VLRs, we constructed large VLR libraries from antigen-stimulated and naïve animals in a novel yeast surface-display vector, with the VLR C-terminally fused to the yeast Flo1p surface anchor. We cloned VLRB binders of lysozyme, β-gal, cholera toxin subunit B, R-phycoerythrin, and B-trisaccharide antigen, with dissociation constants up to the single-digit picomolar range, equivalent to those of high-affinity IgG antibodies. We also isolated from a single lamprey 13 anti-lysozyme VLRA clones with affinities ranging from low nanomolar to mid-picomolar. All of these VLRA clones were closely related in sequence, differing at only 15 variable codon positions along the 244-residue VLR diversity region, which augmented antigen-binding affinity up to 100-fold. Thus, VLRs can provide a protective humoral antipathogen shield. Furthermore, the broad range of nominal antigens that VLRs can specifically bind, and the affinities achieved, indicate a functional parallelism between LRR-based and Ig-based antibodies. VLRs may be useful natural single-chain alternatives to conventional antibodies for biotechnology applications.

DOI： 10.1073/pnas.0904443106

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その他リンク： http://orcid.org/0000-0003-3047-6942

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Nature Structural and Molecular Biology  

Variable lymphocyte receptors (VLRs) are leucine-rich repeat proteins that mediate adaptive immunity in jawless vertebrates. VLRs are fundamentally different from the antibodies of jawed vertebrates, which consist of immunoglobulin (Ig) domains. We determined the structure of an anti-hen egg white lysozyme (HEL) VLR, isolated by yeast display, bound to HEL. The VLR, whose affinity resembles that of IgM antibodies, uses nearly all its concave surface to bind the protein, in addition to a loop that penetrates into the enzyme active site. The VLR-HEL structure combined with sequence analysis revealed an almost perfect match between ligand-contacting positions and positions with highest sequence diversity. Thus, it is likely that we have defined the generalized antigen-binding site of VLRs. We further demonstrated that VLRs can be affinity-matured by 13-fold to affinities as high as those of IgG antibodies, making VLRs potential alternatives to antibodies for biotechnology applications. © 2009 Nature America, Inc. All rights reserved.

DOI： 10.1038/nsmb.1619

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記述言語：英語   出版者・発行元：Elsevier {BV}  

DOI： 10.1016/j.jbiotec.2008.07.537

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出版者・発行元：Informa {UK} Limited  

DOI： 10.1201/9781420006971.ch7

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Journal of Immunology  

Invertebrates display effective innate immune responses for defense against microbial infection. However, the protozoan parasite Perkinsus marinus causes Dermo disease in the eastern oyster Crassostrea virginica and is responsible for catastrophic damage to shellfisheries and the estuarine environment in North America. The infection mechanisms remain unclear, but it is likely that, while filter feeding, the healthy oysters ingest P. marinus trophozoites released to the water column by the infected neighboring individuals. Inside oyster hemocytes, trophozoites resist oxidative killing, proliferate, and spread throughout the host. However, the mechanism(s) for parasite entry into the hemocyte are unknown. In this study, we show that oyster hemocytes recognize P. marinus via a novel galectin (C. virginica galectin (CvGal)) of unique structure. The biological roles of galectins have only been partly elucidated, mostly encompassing embryogenesis and indirect roles in innate and adaptive immunity mediated by the binding to endogenous ligands. CvGal recognized a variety of potential microbial pathogens and unicellular algae, and preferentially, Perkinsus spp. trophozoites. Attachment and spreading of hemocytes to foreign surfaces induced localization of CvGal to the cell periphery, its secretion and binding to the plasma membrane. Exposure of hemocytes to Perkinsus spp. trophozoites enhanced this process further, and their phagocytosis could be partially inhibited by pretreatment of the hemocytes with anti-CvGal Abs. The evidence presented indicates that CvGal facilitates recognition of selected microbes and algae, thereby promoting phagocytosis of both potential infectious challenges and phytoplankton components, and that P. marinus subverts the host's immune/feeding recognition mechanism to passively gain entry into the hemocytes. Copyright © 2007 by The American Association of Immunologists, Inc.

DOI： 10.4049/jimmunol.179.5.3086

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その他リンク： http://orcid.org/0000-0003-3047-6942

記述言語：日本語   出版者・発行元：Advances in Experimental Medicine and Biology  

DOI： 10.1007/978-0-387-71767-8_27

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記述言語：日本語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Springer Nature  

DOI： 10.1111/j.1444-2906.2006.01172.x

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology  

Pufflectin-s, identified in the skin mucus of the fugu Takifugu rubripes, is a novel mannose-specific lectin with similar structure to monocotyledonous plant lectins. In the present study, mutational analysis was used to reveal the mannose-binding sites of pufflectin-s. Putative binding sites were mutated as follows: binding site 1; rPL-D32E (Asp32 → Glu32), rPL-N34S (Asn34 → Ser34) and rPL-V36A (Val 36 → Ala36) whereas binding site 2; rPL-D61E (Asp61 → Glu61), rPL-N63S (Asn63 → Ser63) and rPL-V65A (Val65 → Ala65). All recombinant proteins were expressed in Escherichia coli, purified with two chromatographic steps, and then subjected to mannose-binding assay by affinity chromatography. Recombinant wild-type pufflectin-s (rPL-wt) as well as three mutants with changes in binding site 2 could bind to mannose, in contrast to the three mutants with changes in binding site 1 in which mannose-binding activity was completely lost. These results clearly demonstrate that, at the least, binding site 1 is critical to mannose-binding activity in pufflectin-s. © 2006 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.cbpb.2006.01.005

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Comparative Biochemistry and Physiology - Part D: Genomics and Proteomics  

In earlier work, we identified a novel mannose-specific lectin, termed pufflectin-s, from the skin mucus of torafugu, Takifugu rubripes. We here make a brief review on the lectin. The amino acid sequence of pufflectin-s shares sequence homology with mannose-binding lectins of monocotyledonous plants, and has conserved two of three carbohydrate recognition domains, QDNVY motifs, of these plant lectins. By site-directed mutagenesis, we verified that the QDNVY motif in the N-terminal region was critical to the mannose-binding function of pufflectin-s. RT-PCR and Northern blot analyses indicated that the pufflectin-s gene is expressed in the gill, oral cavity wall, esophagus, and skin. In addition, an isoform, pufflectin-i, which shares 91.4% amino acid identity with pufflectin-s, was isolated from the intestine. Using immunohistochemistry, pufflectin-s could be detected exclusively in the epithelial cells of the skin, gill, oral cavity wall and esophagus, whereas pufflectin-i was observed in both mucous and epithelial cells in the intestine. Nevertheless, mRNAs for both pufflectins were detected only in epithelial cells of these tissues with in situ hybridization. Pufflectin-s agglutinated some bacteria isolated from rearing water and from fish skin. This lectin also bound to a parasite, Heterobothrium okamotoi, suggesting that it may play an important role in the self-defense system of fugu. © 2005 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.cbd.2005.09.005

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記述言語：日本語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Elsevier {BV}  

Two lactose-binding lectins (PFL-1 and -2) were identified in the skin mucus of ponyfish, Leiognathus nuchalis. The molecular masses of PFL-1 and -2 were estimated to be 24 and 30 kDa, respectively. Cloning of the PFL-1 cDNA demonstrated its unique tandem repeat structure composed of two homologous domains with 41.7% internal identity. Furthermore, PFL-1 exhibited homology with l-rhamnose-binding lectins previously purified from the eggs of fish and sea urchins. The N-terminal amino acid sequence of PFL-2 was similar to that of PFL-1, suggesting that this protein is an isotype of PFL-1. The PFL-1 gene was expressed in the skin, an important line of defense against pathogens in fish, but was not expressed in any of the other tissues tested here. PFL-1 is the fourth type of fish skin mucus lectin to be identified, suggesting that different species of fish express different types of lectin in their skin mucus. © 2005 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2005.05.118

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Scopus

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Developmental and Comparative Immunology  

We have previously identified two novel mannose-specific lectins, skin- and intestine-type pufflectins, in the pufferfish, Fugu rubripes [J. Biol. Chem. 278 (2003) 20882]. In the present study, the localization of the lectins and their producing cells were analyzed with antibody and anti-sense probe that recognize both types of pufflectin. Using immunohistochemistry, pufflectins were detected exclusively in epithelial cells in the skin, gills, oral cavity wall and esophagus, whereas in both mucous and epithelial cells in the intestine. Messenger RNAs for pufflectins were detected only in epithelial cells of these tissues with in situ hybridization, suggesting that epithelial cells are able to produce the lectins. Pufflectins are produced and distributed in cells that cover the external and internal body surfaces, which might mean that Fugu have a common immunological system on both surfaces. © 2004 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.dci.2004.06.005

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Scopus

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Developmental and Comparative Immunology  

The characteristics and primary structure of AJL-1, one of the lectins in the skin mucus of the Japanese eel (Anguilla japonica), were examined. This lectin exhibited β-galactoside specific activity in a Ca2+ independent manner. We previously reported that its molecular mass was 16,091Da, although it was approximately 30 kDa as determined by gel filtration, indicating that it is a homodimer having non-covalent bonds. This lectin was composed of 142 amino acid residues having no half-cystinyl residues, and showed homology to members of the galectin family, especially to proto-type galectins. Gene expression of this lectin was detected in skin only, and relative expression was high in an individual that exhibited resistance to infectious disease. AJL-1 showed agglutinating activity against pathogenic bacteria, Streptococcus difficile. This suggests that AJL-1 functions as an important defensive factor at the body surface. © 2003 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.dci.2003.08.006

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Scopus

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology  

Among lectins in the skin mucus of fish, primary structures of four different types of lectin have been determined. Congerin from the conger eel Conger myriaster and AJL-1 from the Japanese eel Anguilla japonica were identified as galectin, characterized by its specific binding to β-galactoside. Eel has additionally a unique lectin, AJL-2, which has a highly conserved sequence of C-type lectins but displays Ca2+- independent activity. This is rational because the lectin exerts its function on the cutaneous surface, which is exposed to a Ca2+ scarce environment when the eel is in fresh water. The third type lectin is pufflectin, a mannose specific lectin in the skin mucus of pufferfish Takifugu rubripes. This lectin showed no sequence similarity with any known animal lectins but, surprisingly, shares sequence homology with mannose-binding lectins of monocotyledonous plants. The fourth lectin was found in the ponyfish Leiognathus nuchalis and exhibits homology with rhamnose-binding lectins known in eggs of some fish species. These lectins, except ponyfish lectin, showed agglutination of certain bacteria. In addition, pufflectin was found to bind to a parasitic trematode, Heterobothrium okamotoi. Taken together, these results demonstrate that skin mucus lectins in fish have wide molecular diversity. © 2003 Elsevier Inc. All rights reserved.

DOI： 10.1016/S1096-4959(03)00178-7

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Journal of Biological Chemistry  

We have characterized pufflectin, a novel mannose-specific lectin, from the skin mucus of the pufferfish, Fugu rubripes. Molecular mass estimations by gel filtration and matrix-assisted laser desorption ionization time-of-flight mass spectrometry and the SDS-PAGE pattern suggest that pufflectin is a homodimer composed of non-covalently associated subunits of 13 kDa. The full-length pufflectin cDNA consists of 527 bp, with 116 amino acid residues deduced from the open reading frame. The amino acid sequence of pufflectin shows no homology with any known animal lectin. Surprisingly, pufflectin shares sequence homology with mannose-binding lectins of monocotyledonous plants and has conserved two of three carbohydrate recognition domains of these plant lectins. The pufflectin gene is expressed in gills, oral cavity wall, esophagus, and skin. In addition, an isoform occurs exclusively in the intestine. Pufflectin differs from mannose-binding lectins purified from the blood plasma of Fugu. Whereas pufflectin did not agglutinate five bacterial species tested, it was demonstrated to bind to the parasitic trematode, Heterobothrium okamotoi. This finding suggests that pufflectin contributes to the parasite-defense system in Fugu.

DOI： 10.1074/jbc.M301038200

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Scopus

記述言語：英語   掲載種別：研究論文（国際会議プロシーディングス）   出版者・発行元：Springer Nature  

DOI： 10.1007/978-4-431-65907-5_33

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Journal of Biological Chemistry  

Two types of lactose-binding lectins, AJL-1 and AJL-2, were purified from the skin mucus extract of the Japanese eel Anguilla japonica by lactose affinity chromatography and subsequent gel filtration. The molecular masses of AJL-1 and AJL-2 were 16,091 and 31,743 Da, respectively. Intact AJL-1 was comprised of two identical 16-kDa subunits having blocked N termini and no disulfide bonds. AJL-2 was a homodimer with disulfide bonds. Based on the N-terminal amino acid sequence of the AJL-2 monomer, the nucleotide sequence of cDNA encoding this lectin was determined by 3′- and 5′-rapid amplification of cDNA ends. The deduced amino acid sequence showed ∼30% homology with C-type lectins, which bind to carbohydrates in a Ca2+-dependent manner. In addition, AJL-2 exhibited highly conserved consensus amino acid residues of the C-type carbohydrate recognition domain, although this lectin showed Ca2+-independent activity. Gene expression of AJL-2 was de. tected only in the skin by Northern blot analysis, and this lectin localization was demonstrated in the club cells by immunohistochemistry. These results indicate that AJL-2 is secreted on the body surface and function as a component of skin mucus. AJL-2 agglutinated Escherichia coli and suppressed its growth, suggesting that this lectin is involved in host defense.

DOI： 10.1074/jbc.M202648200

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：ELSEVIER SCIENCE BV  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：ELSEVIER SCIENCE BV  

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記述言語：日本語   出版者・発行元：Nippon Suisan Gakkaishi (Japanese Edition)  

DOI： 10.2331/suisan.WA2440-7

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記述言語：日本語   出版者・発行元：Nippon Suisan Gakkaishi (Japanese Edition)  

DOI： 10.2331/suisan.WA2440-1

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記述言語：日本語   掲載種別：記事・総説・解説・論説等（学術雑誌）  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：OXFORD UNIV PRESS INC  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：OXFORD UNIV PRESS INC  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：OXFORD UNIV PRESS INC  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：OXFORD UNIV PRESS INC  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：OXFORD UNIV PRESS INC  

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記述言語：日本語   掲載種別：記事・総説・解説・論説等（学術雑誌）  

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記述言語：日本語   掲載種別：記事・総説・解説・論説等（学術雑誌）  

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