Biomedical subjects

I Hirono

Publications and source records attributed to I Hirono.

At least 37 records · Page 2Linked to original sources

PubMed2000

A survey of expressed genes in the leukocytes of Japanese flounder, Paralichthys olivaceus, infected with Hirame rhabdovirus.

We constructed a cDNA library of Japanese flounder, Paralichthys olivaceus, leukocytes that were infected with Hirame rhabdovirus (HRV) in order to analyze some of the genes that are induced and expressed by virus infection in the immune system. Four hundred and fifty-two partial sequences representing 300 cDNA clones were obtained from the 5' and/or 3' ends of inserts derived from the Japanese flounder leukocyte cDNA library. About three-quarters of the 300 cDNA clones (217 clones, 72.3%) represented known genes in the public databases, whereas the remaining 83 (27.7%) of the clones did not show any significant homology with the sequences in the public databases. Clones matching known genes were classified into 12 categories according to their function or distribution. Only 40 (18.4%) of the 217 known genes showed homology with fish genes deposited in the database. Thirty (10%) of the clones, encoding 21 different sequences, and representing several categories, were identified as putative biodefense genes or genes associated with the immune response. Nineteen of the 21 putative biodefense or immune response-related cDNAs have not been previously reported in fish genes or cDNAs.

Amino Acid Sequence↗

PubMed2000

Cloning and analysis of expression of Mx cDNA in Japanese flounder, Paralichthys olivaceus.

Myxovirus resistance (Mx) is an interferon (IFN)-induced protein that specifically prevents growth, in vivo and in vitro, of certain classes of virus. We isolated the Mx cDNA from a leukocyte cDNA library obtained from a Japanese flounder infected with Hirame rhabdovirus (HRV). The Mx cDNA was found to contain 2385 bp coding for 620 amino acids. The amino acid sequence had approximately 51 and 78% identities to the sequences of human and rainbow trout Mx cDNAs, respectively. We propose that the amino acid sequence encoded by this cDNA clone contains a tripartite guanosine-5'-triphosphate (GTP)-binding motif, GXXXSGKS/T, DXXG and T/NKXD, and a signature of the dynamin family, LPRG(S/K)GIVTR, which are conserved in all IFN-induced Mx proteins of vertebrates. In addition, the C-terminal region of the Japanese flounder Mx protein, like those of other Mx proteins, contains a highly conserved leucine zipper motif. Japanese flounder infected with the HRV showed increased expression of Mx mRNA in the leukocytes starting at 48 h and peaking at 72 h. An RT-PCR (reverse transcription-polymerase chain-reaction) analysis of Mx mRNA from a healthy fish showed that the Mx mRNA was predominantly expressed in the kidney, spleen, intestine, brain, peritoneal cavity fluid and gill, and only minimally expressed in leukocytes, liver, muscle and mucus.

Amino Acid Sequence↗

PubMed1999

Identification of genes in a KG- phenotype of Lactococcus garvieae, a fish pathogenic bacterium, whose proteins react with antiKG- rabbit serum.

Five different clones (SA1B05, SA1B10, SA2F01, SA8A11 and SA9H10) were isolated from the gene library of the Lactococcus garvieae SA8201 (KG-) strain by immunological screening using rabbit serum against L. garvieae (KG-) phenotype cells. A Western blot analysis indicated that the molecular sizes of immunologically detected proteins of SA1B05, SA1B10, SA2F01, SA8A11 and SA9H10, which were fused with LacZ protein, were 25, 30, 28, 26 and 13 kDa, respectively. The amino acid sequences of the immunologically detected proteins of SA1B05, SA1B10, SA2F01 and SA8A11 were homologous to a processing protease of Bacillus subtilis (36.6%), dihydropteroate synthase of Escherichia coli (34.6%), trigger factor of B. subtilis (45.8%) and N-acetylglucosamine-6-phosphate deacetylase of Vibrio furnissii (37.1%), respectively. There was no significant homologous sequence of SA9H10 in DDBJ/EMBL/GenBank and SwissProt. We cloned and sequenced a longer DNA fragment (SA9H10L) of SA9H10 from the gene library. The predicted amino acid sequence of this clone was weak homology to M protein of Streptococcus pyogenes (22.7%). Five genes were specifically expressed in the KG- phenotype strains. However, SA8A11 and SA9H10 was expressed in the mutated strain SA8201-TTC, whose serological phenotype was changed from KG- to KG+ by 2,3,5-triphenyltetrazolium chloride.

Amino Acid Sequence↗

PubMed1999

Sequences of 596 cDNA Clones (565,977 bp) of Japanese Flounder (Paralichthys olivaceus) Leukocytes Infected with Hirame Rhabdovirus.

: We have partially sequenced 785 sequences of 596 independent complementary DNA clones isolated from a cDNA library of Japanese flounder leukocytes infected with hirame rhabdovirus. These sequences consist of a total of 565,977 base pairs. The average size of the sequenced lengths was 721 bp. Of 596 clones, 386 (64.8%) were identified as previously reported genes by the BLASTN and BLASTX programs. About 30% of the identified clones could be recognized by only the BLASTX program. A total of 251 distinct genes were identified, and 181 of these genes are the first such genes reported from the teleostei. Approximately 27% of the identified Japanese flounder genes appear to be associated with cell division, cell structure or motility, and basic energy metabolism, 29% with gene or protein expression, and 17% with cell signaling, cell communication, and cell or organism defense. The most frequently identified expressed sequence tags of leukocytes of Japanese flounder were gelatinase b and ribosomal protein L23, which both had 1.34% prevalence.

Journal Article↗

PubMed1999

Molecular analysis of complement component C8beta and C9 cDNAs of Japanese flounder, Paralichthys olivaceus.

The amino acid sequences of the human terminal complement components show extensive structural similarity to each other. In this study the C8beta and C9 cDNAs of Japanese flounder, Paralichthys olivaceus, were cloned and analyzed. The derived deduced amino acid sequences of the two terminal components were homologous to those of humans, in that the sequences of both species contained LDL receptor, EGF precursor, and two thrombospondin domains. Japanese flounder C9 was found to have a second thrombospondin region in the C-terminus, similar to that reported for rainbow trout and pufferfish. Moreover, these two complement component cDNAs of Japanese flounder had partial similarity to human perforin. These findings show that Japanese flounder C8beta and C9 have similar structures, which supports the hypothesis that the terminal complement genes originated from the same ancestral gene. Collectively, these features emphasize the strong similarity among the members of the terminal complement family.

Amino Acid Sequence↗

PubMed1998

Rapid diagnosis of red sea bream iridovirus infection using the polymerase chain reaction.

A simple and sensitive polymerase chain reaction (PCR) based assay is described for detection of the red sea bream iridovirus (RSIV) in infected fish. The assay involves amplification of a portion of the ribonucleotide reductase small subunit (RNRS) gene of the virus from DNA isolated from the spleen. The system was tested on red sea bream following an experimental infection. In our infection model, disease signs first became apparent 5 to 6 d post-infection, and mortality commenced at Day 6 and reached 90% by Day 9. No amplified product was detected from fish at 1 or 2 d post-infection, but 3 of 5 fish tested positive at Day 3, and all fish tested positive at Days 5 and 8. Thus, infection could be detected prior to the appearance of overt symptoms. This PCR method should be of considerable value for aquaculture to detect RSIV infection.

Animals↗

PubMed1998

Note: molecular cloning of chitinase genes from Vibrio anguillarum and V. parahaemolyticus.

Chitinase genes from Vibrio anguillarum KV9001 and V. parahaemolyticus ATCC17802 were cloned into Escherichia coli. Open reading frames of chitinase genes from V. anguillarum (vac) and V. parahaemolyticus (vpc) are 1755 bp and 1890 bp, respectively. The deduced amino acid sequences of these genes have 71.6% identity. There are two consensus sequence regions in the VAC and VPC proteins. The vac gene was highly prevalent in V. anguillarum, and the DNA probe of the vac gene hybridized to V. alginolyticus and Beneckea proteolytica DNA. The DNA probe of the upc gene hybridized to V. alginolyticus, V. harveyi and V. ordalii DNA.

Amino Acid Sequence↗

PubMed1998

DNA sequencing of the gene encoding Salmonella typhimurium-derived T-cell inhibitor (STI) and characterization of the gene product, cloned STI.

In a previous study, we found a novel protein which inhibited T-cell responsiveness to interleukin-2 (IL-2) in Salmonella typhimurium and called it S. typhimurium-derived T-cell inhibitor (STI). In this study, we analyzed the DNA sequence of the gene encoding STI. The STI gene was cloned into a plasmid vector, pUC118, and expressed in Escherichia coli JM109. Like native STI, the cloned STI inhibited IL-2-dependent CTLL-2 cell growth. Furthermore, this growth inhibition involved down-regulation of IL-2 receptor expression. These results indicate that the cloned STI expressed in E. coli was identical to native STI. Sequencing revealed that the STI gene contained an open reading frame of 2298 base pairs encoding a precursor form of 765 amino acid residues (molecular mass 83605) that is processed into a mature form of 745 amino acid residues with molecular mass 81 548. Homology analysis revealed that its amino acid sequence was highly homologous with that of the beta-glucosidase of E. coli K-12. We designated the gene encoding STI sti.

Amino Acid Sequence↗

PubMed1998

Structure and expression of activin genes in rainbow trout.

Activins are dimeric members of the transforming growth factor-beta (TGF-beta) superfamily. By using the polymerase chain reaction (PCR), we have cloned and sequenced activin beta A and beta B genes encoding the mature region of the peptides from the rainbow trout genomic DNA. Two forms of beta A and a single form of beta B-subunits were found. There is high identity with mammalian counterparts; the two rainbow trout beta A-subunits have more than 75% nucleotide sequence identity with the human beta A-subunit, and the beta B-subunit had 82% sequence identity with the human beta B-subunit. Expression of rainbow trout activin genes was examined by reverse transcription-PCR (RT-PCR). The major expression tissue of rainbow trout activin was ovary and brain at the messenger RNA level, and the major expression subunit of rainbow trout activin was the beta B subunit.

Activins↗

PubMed1998

Molecular cloning of a novel interferon regulatory factor in Japanese flounder, Paralichthys olivaceus.

A complementary DNA library was constructed in lambda ZAP II using messenger RNA from the leukocytes of some heterocloned Japanese flounder, Paralichthys olivaceus, that had been artificially infected with Hirame rhabdovirus (HRV). A cloned flounder interferon regulatory factor (designated fIRF) cDNA was found to be 1746 bp in length, with an open reading frame of 297 amino acids. The overall amino acid sequence of fIRF had approximately 40% identity with the previously reported avian and mammalian IRF-1s and IRF-2s. The fIRF sequence was most similar to that recorded for the chicken IRF-1. Amino acid sequence identities between the DNA-binding domain of the fIRF and that of both chicken IRF-1 and chicken IRF-2 were 72.3%. The DNA-binding domain of fIRF contained the repeated tryptophan motif that is characteristic of members of the IRF family. The mRNA of fIRF was detected in various tissues by reverse transcription-polymerase chain reaction (RT-PCR). The fIRF was transcribed mainly in the intestine, ovary, muscle, liver, heart and spleen, while it was minimally transcribed in the brain and kidney. When Japanese flounder were injected with HRV, the relative expression of fIRF mRNA was found to increase and peak 3 days after injection. The quantities of the fIRF mRNA increased to levels that were 7.5-fold higher than those of noninjected fish. In addition, when Japanese flounder were injected with Edwardsiella tarda, the expression of fIRF mRNA showed increases 2, 3, and 4 days after injection. The quantities of the fIRF mRNA on those days represented approximately 6-, 15-, and 14-fold increases, respectively, over the levels in noninjected fish.

Amino Acid Sequence↗

PubMed1998

Molecular cloning and evolution of transferrin cDNAs in salmonids.

Transferrin complementary DNAs were cloned from the livers of seven species in three genera of salmonids (kokanee salmon Oncorhynchus nerka, amago salmon Oncorhynchus masou ishikawa, masu salmon Oncorhynchus masou masou, Japanese char Salvelinus pluvius, brook trout Salvelinus fontinalis, lake trout Salvelinus namaycush, and brown trout Salmo trutta) subsequent to polymerase chain reaction amplification with primers derived from conserved regions of transferrin cDNA sequences. The transferrin cDNAs of the seven species of salmonids had sizes of 2.2 to 2.4 kb and encoded an open reading frame consisting of 691 amino acids with a putative signal peptide of 18 amino acids. The alignment of salmonid transferrin cDNAs showed a duplicated structure and conserved anion-binding residues, iron-binding residues, and cysteine residues for disulfide bridges. The deduced amino acid sequences of the seven salmonid transferrin cDNAs share 85% to 99% homology. A phylogenetic tree of amino acid sequences of transferrin cDNAs from salmonids showed that the relationship among the three genera of salmonids (Oncorhynchus, Salvelinus, and Salmo) is well correlated with that derived from classic morphologic and genetic analyses.

Amino Acid Sequence↗

PubMed1997

Identification of major antigenic proteins of Pasteurella piscicida.

Two different antigenic protein-coding clones (PPA1 and PPA2) were isolated using anti-Pasteurella piscicida rabbit serum from a genomic DNA library of P. piscicida strain KP9038. The PPA1 and PPA2 expressed 7 kDa and 45 kDa proteins in Escherichia coli, respectively, and the molecular sizes of these expressed proteins are the same as these of the major antigenic proteins of P. piscicida. PPA1 encodes a protein of 83 amino acids residues, which is similar to the bacterial lipoprotein. Comparison of the predicted amino acid sequence of the PPA1-encoded 7 kDa protein of P. piscicida with previously reported bacterial lipoprotein sequence data revealed that it shares about 40% amino acid sequence identity. PPA2 has two large open reading frame (ORFs). The larger ORF (encoding 452 amino acid residues) encodes a homolog of DegQ protease, and the smaller ORF (371 amino acid residues) encodes a homolog of DegS protease. The antibodies reacted with the larger ORF-encoded 45 kDa DegQ homolog protein. The DegQ and DegS homolog proteins contain an export signal and a serine protease active site. The structural features of the PPA2-coding locus are similar to those of the loci in E. coli for the degQ and degS serine protease genes. A sequence in the 3' non-coding region of Vibrio hollisae thermostable hemolysin gene that is highly homologous with a similar located sequence in the Pseudomonas putida p-cresol methylhydroxylase gene is also found in the 3' non-coding region of the degS homolog gene of the PPA2.

Amino Acid Sequence↗

PubMed1997

Iron-regulated haemolysin gene from Edwardsiella tarda.

We have cloned and sequenced the haemolysin gene locus from Edwardsiella tarda (ETH). This region encoded two open reading frames, designated ethA and ethB. ethA is the haemolysin gene consisting of 4782bp encoding a product of 165.3 kDa and ethB is an activation/secretion protein gene of 1677bp that encodes a product of 61.9 kDa. There were two putative ferric uptake regulator (Fur) binding sites on the 5' upstream region of the ethB gene overlapping the promoter region and ribosome-binding site. The haemolysin produced by the cloned gene was secreted by Escherichia coli. The deduced amino acid sequences of the ethA and ethB genes were found to be homologous to those of the haemolysin and activation/secretion proteins of Haemophilus ducreyi, Proteus mirabilis, and Serratia marcescens. E. coli carrying the ethA gene but not the ethB gene completely lost haemolytic activity, although the ethA gene was transcribed. The protein expressed by E. coli carrying a recombinant plasmid which encoded the ethA gene had haemagglutination activity. The EthB protein was necessary for activation of EthA protein (haemolysin). The ethA and ethB genes were very prevalent in haemolytic E. tarda strains isolated from diseased fish. Transcription of the ethB gene was regulated by iron. The ethA and ethB genes were transcribed independently.

Amino Acid Sequence↗

PubMed1997

Demonstration of ras and p53 gene mutations in carcinomas in the forestomach and intestine and soft tissue sarcomas induced by N-methyl-N-nitrosourea in the rat.

The presence of ras family and p53 gene mutations in rat forestomach, intestine and liver tumors and soft tissue sarcomas induced by N-methyl-N-nitrosourea (MNU) was examined using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) followed by direct sequencing analysis. In the forestomach squamous cell carcinomas (SCC), Ha-ras and p53 mutations were detected in 2 (40%) and 4 (80%) of 5 cases, respectively. The figures for Ki-ras and p53 gene mutations in adenocarcinomas of the large and small intestines were 3 (18.8%) and 5 (31.3%) of 16 cases. Soft tissue sarcomas in different sites were found to have mutations of Ki-ras in 7 (23.3%) and of p53 in 9 (30%) of 30 cases. One forestomach SCC and 2 soft tissue sarcomas had double p53 mutations in different exons. Single cases of forestomach SCC and intestinal adenocarcinoma had mutations in both Ki-ras and p53 genes. No mutations were found in counterpart benign tumors or hepatocellular adenomas. The p53 mutation spectrum revealed preferential clustering within exon 8 for the forestomach SCCs, and exons 5 and 8 for the intestinal adenocarcinomas, whereas the distribution was evenly spread through exons 5 to 8 in soft tissue sarcomas. All the detected ras or p53 mutations were G:C to A:T transitions. These results indicate firstly that specific Ki-ras, Ha-ras and p53 gene mutations in MNU-induced lesions are related to particular alkylation sites (G:C to A:T transitions) and secondly, although not essential, Ki-ras, Ha-ras or p53 gene mutations may be involved in the progression stage of forestomach, intestine and soft tissue neoplasms induced by MNU.

Adenocarcinoma↗

PubMed1997

Characterization and expression of c-type lysozyme cDNA from Japanese flounder (Paralichthys olivaceus).

Lysozyme is a widely distributed enzyme located in the serum, skin mucus, and other organs of fish, which is responsible for catalyzing the hydrolysis of the cell walls of most bacteria. A c-type of lysozyme cDNA was cloned from a kidney cDNA library of the Japanese flounder (Paralichthys olivaceus). The cDNAs consisted of 612 bp, which coded for 143 amino acid residues. The deduced amino acid sequence of Japanese flounder c-type lysozyme possessed 72.9%, 57.4%, and 65.4% identities with rainbow trout, chicken, and human c-type lysozymes, respectively. Comparison of the c-type lysozymes showed that the catalytic residues, the residues binding to sugar chains, and cysteine residues were completely conserved. Northern blot analysis indicated that the c-type lysozyme gene is apparently transcribed in the head kidney, posterior kidney, spleen, brain, and ovary of healthy flounder. When flounder were experimentally infected with Edwardsiella tarda, quantities of the c-type lysozyme mRNA increased in the head kidney, spleen, and ovary of the flounder.

Amino Acid Sequence↗

PubMed1997

Expressed sequence tags of medaka (Oryzias latipes) liver mRNA.

A medaka liver cDNA library was constructed in lambda ZAPII. The number of clones in this library is approximately 5 x 10(6). Three hundred sixty-one clones were randomly selected and, of these, 33 clones with over 1 kb were sequenced. These sequences were compared with GenBank and the dbEST (Re.96.0). Twenty-five clones of these 33 clones encoded 18 different genes and 2 different expressed sequence tags (ESTs). Sequences of 10 of the 18 clones had not previously been reported in fish. The codon usage of medaka genes is similar to that of Xenopus, mouse, and human genes, but not similar to that of yeast and Escherichia coli genes.

Animals↗

PubMed1997

Cloning and characterization of transferrin cDNA and rapid detection of transferrin gene polymorphism in rainbow trout (Oncorhynchus mykiss).

A cDNA clone of rainbow trout (Oncorhynchus mykiss) transferrin was obtained from a liver cDNA library. The 2537-bp cDNA sequence contained an open reading frame encoding 691 amino acids and the 5' and 3' noncoding regions. The amino acid sequences at the iron-binding sites and the two N-linked glycosylation sites, and the cysteine residues were consistent with known, conserved vertebrate transferrin cDNA sequences. Single N-linked glycosylation sites existed on the N- and C-lobe. The deduced amino acid sequence of the rainbow trout transferrin cDNA had 92.9% identities with transferrin of coho salmon (Oncorhynchus kisutch); 85%, Atlantic salmon (Salmo salar); 67.3%, medaka (Oryzias latipes); 61.3% Atlantic cod (Gadus morhua); and 59.7%, Japanese flounder (Paralichthys olivaceus). The long and accurate polymerase chain reaction (LA-PCR) was used to amplify approximately 6.5 kb of the transferrin gene from rainbow trout genomic DNA. Restriction fragment length polymorphisms (RFLPs) of the LA-PCR products revealed three digestion patterns in 22 samples.

Amino Acid Sequence↗

PubMed1997

A survey of expressed genes in Japanese flounder (Paralichthys olivaceus) liver and spleen.

Expressed sequence tags (ESTs) were obtained from Japanese flounder (Paralichthys olivaceus). We present the results of single-pass sequencing of 493 ESTs from 350 clones from liver cDNA and 57 ESTs from 41 clones from spleen cDNA. Sequences of the cDNA clones were compared with sequences in the GenBank database. Two hundred and two clones (51.7%) appeared to be completely unknown and are likely to represent newly described genes, whereas 189 clones (48.3%) were identified based on matches to sequences in the databases. Three of the unidentified sequences were isolated from both the liver and spleen cDNA libraries. However, there were no identical sequences between liver and spleen clones.

Animals↗