Establishment of a novel clonal GFP-expressing transgenic ginbuna crucian carp.

The clonal triploid ginbuna crucian carp Carassius auratus langsdorfii, a naturally occurring gynogenetic fish, is suitable for cell transplantation studies to reveal the roles of stem cells and immune cells. To ensure long-term traceability of donor cells within recipient fish, we have established a transgenic ginbuna line that expresses green fluorescent protein (GFP). The Xenopus laevis ef1a promoter was introduced for regulating GFP expression.

Characterization of fish-specific IFNγ-related binding with a unique receptor complex and signaling through a novel pathway.

Unlike mammals, fish express two type II interferons, IFNγ and fish-specific IFNγ (IFNγ-related or IFNγrel). We previously reported the presence of two IFNγrel genes, IFNγrel 1 and IFNγrel 2, which exhibit potent antiviral activity in the Ginbuna crucian carp, Carassius auratus langsdorfii. We also found that IFNγrel 1 increased allograft rejection; however, the IFNγrel 1 receptor(s) and signaling pathways underlying this process have not yet been elucidated.

Alteration of hemoglobin ß gene expression in mucosal tissues of Japanese flounder, , in response to heat stress, infection, and immunostimulants administration.

Hemoglobin beta (Hbß) is a heme-binding protein capable of oxygen delivery. The oligopeptides derived from Hbβ in fish mucus are active against a variety of gram-negative bacteria and protozoa. To gain information on the physiological and immunological roles of Hbβ in the mucosal tissues of fish, we analyzed changes in Hbß gene expression levels in the epidermis, gills, and intestine of Japanese flounder, , in response to heat stress, infection, and trial feeding of immunostimulants, high-concentration ascorbic acid (AsA) or lactoferrin (LF).

Plastidial starch phosphorylase is highly associated with starch accumulation process in developing squash (Cucurbita sp.) fruit.

We investigated changes in starch content and starch metabolic enzyme activities in developing and postharvest squash of distinct species, Cucurbita maxima and Cucurbita moschata, which accumulate high and low levels of starch, respectively. The total activity of starch phosphorylase in developing fruits significantly correlated (r = 0.99) to the amount of starch among Cucurbita species (C.

A designed cell-penetrating human SOCS2 protein suppresses GH-dependent cancer cell proliferation.

Suppressor of cytokine signaling (SOCS) 2, a negative regulator of growth hormone (GH) and insulin-like growth factor 1 (IGF-1), which is associated with acromegaly and cancers, is a promising candidate molecule for treating various diseases. To facilitate its use in protein therapy, we designed and constructed a human SOCS2 protein containing a membrane-permeable peptide sequence and expressed it in an Escherichia coli system. The partially purified recombinant protein was effectively delivered into several cancer cell lines and inhibited cell growth.

Sequential steps of macroautophagy and chaperone-mediated autophagy are involved in the irreversible process of posterior silk gland histolysis during metamorphosis of Bombyx mori.

To elucidate the degradation process of the posterior silk gland during metamorphosis of the silkworm ITALIC! Bombyx mori, tissues collected on the 6th day after entering the 5th instar (V6), prior to spinning (PS), during spinning (SP) and after cocoon formation (CO) were used to analyze macroautophagy, chaperone-mediated autophagy (CMA) and the adenosine triphosphate (ATP)-dependent ubiquitin proteasome. Immediately after entering metamorphosis stage PS, the levels of ATP and phosphorylated p70S6 kinase protein decreased spontaneously and continued to decline at SP, followed by a notable restoration at CO. In contrast, phosphorylated AMP-activated protein kinase α (AMPKα) showed increases at SP and CO.

Purification and characterization of a fish granzymeA involved in cell-mediated immunity.

Granzymes are serine proteases involved in the induction of cell death against non-self cells. The enzymes differ in their primary substrate specificity and have one of four hydrolysis activities: tryptase, Asp-ase, Met-ase and chymase. Although granzyme genes have been isolated from several fishes, evidence for their involvement in cytotoxicity has not yet been reported.

Detecting Dengue Virus Nonstructural Protein 1 (NS1) in Urine Samples Using ELISA for the Diagnosis of Dengue Virus Infection.

Dengue virus (DENV) infection is a serious global health threat. For the surveillance and control of dengue, there is a need for robust diagnostic tools that are relatively easy to use and reliable in various clinical settings. We investigated the applicability of NS1 antigen detection in urine samples for the diagnosis of DENV.

Identification of a novel fish granzyme involved in cell-mediated immunity.

Granzymes (Gzms) are serine proteases released from cytoplasmic granules within cytotoxic T lymphocytes and natural killer (NK) cells. Gzms induce apoptosis within virus-infected and transformed cells. In fish as well as mammals, Gzms appear to play a major role in inducing target cell death.

Peculiar monomeric interferon gammas, IFNγrel 1 and IFNγrel 2, in ginbuna crucian carp.

The existence of fish-specific isoforms of interferon (IFN)γ, known as IFNγ-related (IFNγrel), has been reported in several fish species. However, comparisons with deduced amino acid sequences of known IFNγrels among several fish species have indicated significant differences at the C-terminus basic amino acid continuous sequences, which indicate the existence of multiple IFNγrel isoforms. Two distinct cDNAs, encoding two IFNγrels, ifngrel 1 and ifngrel 2, were cloned from ginbuna crucian carp (Carassius auratus langsdorfii).

Heterologous expression of tomato glycoside hydrolase family 3 α-L-arabinofuranosidase/β-xylosidases in tobacco suspension cultured cells and synergic action of a family 51 isozyme under antisense suppression of the enzyme.

Four cDNA clones (SlArf/Xyl1-4) encoding α-l-arabinofuranosidase/β-xylosidase belonging to glycoside hydrolase family 3 were obtained from tomato (Solanum lycopersicum) fruit. SlArf/Xyl1 was expressed in various organs. Its level was particularly high in flower and leaves but low in fruit.

Identification and characterization of the short variable region of the Japanese encephalitis virus 3' NTR.

Since the 1980s, the Japanese encephalitis virus (JEV) variants with slightly short variable regions (VR) of the 3' non-translated region (NTR) have been found; however, the implications of these short VR remain unclear. We recently identified two novel types of short VR (5 and 9 nt shorter than that of major group of genotype I JEV strains) of genotype I JEV isolates. To elucidate the impact of these short VR on the replication and virulence of JEV, we generated five recombinant JEV viruses: M41-d5 and M41-d9 have deletions in the VR that correspond to those observed in some recent JEV isolates, M41-d5d9 has both the 5- and 9-nt deletions in the VR, M41-d27 has a large deletion that encompasses both the 5- and 9-nt deletion regions, and M41-a13 has a 13-nt sequence insertion of the genotype III JEV strain Beijing-1 into the parent genotype I JEV strain Mie/41/2002 genome.

Antiviral protection mechanisms mediated by ginbuna crucian carp interferon gamma isoforms 1 and 2 through two distinct interferon gamma-receptors.

Fish genomes possess three type II interferon (IFN) genes, ifnγ1, ifnγ2 and ifnγ-related (ifnγrel). The IFNγ-dependent STAT signalling pathway found in humans and mice had not been characterized in fish previously. To identify the antiviral functions and signalling pathways of the type II IFN system in fish, we purified the ifnγ1, ifnγ2 and ifnγrel proteins of ginbuna crucian carp expressed in bacteria and found them to elicit high antiviral activities against crucian carp hematopoietic necrosis virus.

Evaluating antigen-specific cytotoxicity of CD8+ T cells in fish by granzyme B-like activity.

Granzyme B plays an important role in granule-mediated apoptosis by CTL. It is a well characterized component of the cytolytic machinery in mammals and a candidate for the evaluation of cytotoxic activity of CTL as an alternative to conventional cytotoxicity assay. In this study, we examined the effects of granzyme inhibitors to assess the characteristics of fish granzymes in terms of substrate specificity and the involvement of granzyme B-like in the cytotoxic response.

Diallyl trisulfide suppresses the proliferation and induces apoptosis of human colon cancer cells through oxidative modification of beta-tubulin.

Allyl sulfides are characteristic flavor components obtained from garlic. These sulfides are thought to be responsible for their epidemiologically proven anticancer effect on garlic eaters. This study was aimed at clarifying the molecular basis of this anticancer effect of garlic by using human colon cancer cell lines HCT-15 and DLD-1.