Genomic characterization and expression profiles upon bacterial infection of a novel cystatin B homologue from disk abalone (Haliotis discus discus)
► Complete genomic sequence of cystatin B from disk abalone was identified. ► AbCyt B mRNA was highly expressed in hemocytes, gills, mantle, and digestive tract. ► Bacterial infection caused significant upregulation of AbCyt B expression in gills. ► Recombinant AbCyt B showed strong in vitro papain...
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| Veröffentlicht in: | Developmental and comparative immunology 2012-12, Vol.38 (4), p.495-504 |
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| creator | Premachandra, H.K.A. Wan, Qiang Elvitigala, Don Anushka Sandaruwan De Zoysa, Mahanama Choi, Cheol Young Whang, Ilson Lee, Jehee |
| description | ► Complete genomic sequence of cystatin B from disk abalone was identified. ► AbCyt B mRNA was highly expressed in hemocytes, gills, mantle, and digestive tract. ► Bacterial infection caused significant upregulation of AbCyt B expression in gills. ► Recombinant AbCyt B showed strong in vitro papain inhibitory activity.
Cystatins are a large family of cysteine proteinase inhibitors which are involved in diverse biological and pathological processes. In the present study, we identified a gene related to cystatin superfamily, AbCyt B, from disk abalone Haliotis discus discus by expressed sequence tag (EST) analysis and BAC library screening. The complete cDNA sequence of AbCyt B is comprised of 1967 nucleotides with a 306bp open reading frame (ORF) encoding for 101 amino acids. The amino acid sequence consists of a single cystatin-like domain, which has a cysteine proteinase inhibitor signature, a conserved Gly in N-terminal region, QVVAG motif and a variant of PW motif. No signal peptide, disulfide bonds or carbohydrate side chains were identified. Analysis of deduced amino acid sequence revealed that AbCyt B shares up to 44.7% identity and 65.7% similarity with the cystatin B genes from other organisms. The genomic sequence of AbCyt B is approximately 8.4Kb, consisting of three exons and two introns. Phylogenetic tree analysis showed that AbCyt B was closely related to the cystatin B from pacific oyster (Crassostrea gigas) under the family 1.Functional analysis of recombinant AbCyt B protein exhibited inhibitory activity against the papain, with almost 84% inhibition at a concentration of 3.5μmol/L. In tissue expression analysis, AbCyt B transcripts were expressed abundantly in the hemocyte, gill, mantle, and digestive tract, while weakly in muscle, testis, and hepatopancreas. After the immune challenge with Vibrio parahemolyticus, the AbCyt B showed significant (P |
| doi_str_mv | 10.1016/j.dci.2012.06.010 |
| format | Article |
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Cystatins are a large family of cysteine proteinase inhibitors which are involved in diverse biological and pathological processes. In the present study, we identified a gene related to cystatin superfamily, AbCyt B, from disk abalone Haliotis discus discus by expressed sequence tag (EST) analysis and BAC library screening. The complete cDNA sequence of AbCyt B is comprised of 1967 nucleotides with a 306bp open reading frame (ORF) encoding for 101 amino acids. The amino acid sequence consists of a single cystatin-like domain, which has a cysteine proteinase inhibitor signature, a conserved Gly in N-terminal region, QVVAG motif and a variant of PW motif. No signal peptide, disulfide bonds or carbohydrate side chains were identified. Analysis of deduced amino acid sequence revealed that AbCyt B shares up to 44.7% identity and 65.7% similarity with the cystatin B genes from other organisms. The genomic sequence of AbCyt B is approximately 8.4Kb, consisting of three exons and two introns. Phylogenetic tree analysis showed that AbCyt B was closely related to the cystatin B from pacific oyster (Crassostrea gigas) under the family 1.Functional analysis of recombinant AbCyt B protein exhibited inhibitory activity against the papain, with almost 84% inhibition at a concentration of 3.5μmol/L. In tissue expression analysis, AbCyt B transcripts were expressed abundantly in the hemocyte, gill, mantle, and digestive tract, while weakly in muscle, testis, and hepatopancreas. After the immune challenge with Vibrio parahemolyticus, the AbCyt B showed significant (P<0.05) up-regulation of relative mRNA expression in gill and hemocytes at 24 and 6h of post infection, respectively. These results collectively suggest that AbCyst B is a potent inhibitor of cysteine proteinases and is also potentially involved in immune responses against invading bacterial pathogens in abalone.</description><identifier>ISSN: 0145-305X</identifier><identifier>EISSN: 1879-0089</identifier><identifier>DOI: 10.1016/j.dci.2012.06.010</identifier><identifier>PMID: 22878425</identifier><language>eng</language><publisher>United States: Elsevier Ltd</publisher><subject>abalone ; Amino Acid Sequence ; amino acids ; Animals ; bacterial artificial chromosomes ; Bacterial challenge ; bacterial infections ; Base Sequence ; Cloning, Molecular ; complementary DNA ; Computer Simulation ; Crassostrea gigas ; Cystatin B ; Cystatin B - chemistry ; Cystatin B - genetics ; Cystatin B - metabolism ; cystatins ; Cysteine protease inhibitors ; digestive tract ; Disk abalone ; disulfide bonds ; exons ; expressed sequence tags ; Gastropoda - genetics ; Gastropoda - metabolism ; Gastropoda - microbiology ; gene expression ; Gene Expression Regulation ; genes ; Genome ; Haliotis discus ; hemocytes ; Hemocytes - metabolism ; hepatopancreas ; Host-Pathogen Interactions ; immune response ; infection ; introns ; Models, Molecular ; Molecular Sequence Data ; muscles ; nucleotides ; open reading frames ; Organ Specificity ; papain ; Papain - antagonists & inhibitors ; Papain - chemistry ; Papain inhibition ; pathogens ; Phylogeny ; Proteolysis ; screening ; Sequence Analysis, DNA ; Sequence Homology, Amino Acid ; signal peptide ; Structural Homology, Protein ; testes ; Transcriptional analysis ; Vibrio parahaemolyticus ; Vibrio parahaemolyticus - physiology</subject><ispartof>Developmental and comparative immunology, 2012-12, Vol.38 (4), p.495-504</ispartof><rights>2012 Elsevier Ltd</rights><rights>Copyright © 2012 Elsevier Ltd. All rights reserved.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c480t-896edd578b19bf52d9e1d849265f9db3dd5e550b7ab20ca61367c2dc4f7a8bc73</citedby><cites>FETCH-LOGICAL-c480t-896edd578b19bf52d9e1d849265f9db3dd5e550b7ab20ca61367c2dc4f7a8bc73</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://www.sciencedirect.com/science/article/pii/S0145305X12001553$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,776,780,3537,27901,27902,65306</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/22878425$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Premachandra, H.K.A.</creatorcontrib><creatorcontrib>Wan, Qiang</creatorcontrib><creatorcontrib>Elvitigala, Don Anushka Sandaruwan</creatorcontrib><creatorcontrib>De Zoysa, Mahanama</creatorcontrib><creatorcontrib>Choi, Cheol Young</creatorcontrib><creatorcontrib>Whang, Ilson</creatorcontrib><creatorcontrib>Lee, Jehee</creatorcontrib><title>Genomic characterization and expression profiles upon bacterial infection of a novel cystatin B homologue from disk abalone (Haliotis discus discus)</title><title>Developmental and comparative immunology</title><addtitle>Dev Comp Immunol</addtitle><description>► Complete genomic sequence of cystatin B from disk abalone was identified. ► AbCyt B mRNA was highly expressed in hemocytes, gills, mantle, and digestive tract. ► Bacterial infection caused significant upregulation of AbCyt B expression in gills. ► Recombinant AbCyt B showed strong in vitro papain inhibitory activity.
Cystatins are a large family of cysteine proteinase inhibitors which are involved in diverse biological and pathological processes. In the present study, we identified a gene related to cystatin superfamily, AbCyt B, from disk abalone Haliotis discus discus by expressed sequence tag (EST) analysis and BAC library screening. The complete cDNA sequence of AbCyt B is comprised of 1967 nucleotides with a 306bp open reading frame (ORF) encoding for 101 amino acids. The amino acid sequence consists of a single cystatin-like domain, which has a cysteine proteinase inhibitor signature, a conserved Gly in N-terminal region, QVVAG motif and a variant of PW motif. No signal peptide, disulfide bonds or carbohydrate side chains were identified. Analysis of deduced amino acid sequence revealed that AbCyt B shares up to 44.7% identity and 65.7% similarity with the cystatin B genes from other organisms. The genomic sequence of AbCyt B is approximately 8.4Kb, consisting of three exons and two introns. Phylogenetic tree analysis showed that AbCyt B was closely related to the cystatin B from pacific oyster (Crassostrea gigas) under the family 1.Functional analysis of recombinant AbCyt B protein exhibited inhibitory activity against the papain, with almost 84% inhibition at a concentration of 3.5μmol/L. In tissue expression analysis, AbCyt B transcripts were expressed abundantly in the hemocyte, gill, mantle, and digestive tract, while weakly in muscle, testis, and hepatopancreas. After the immune challenge with Vibrio parahemolyticus, the AbCyt B showed significant (P<0.05) up-regulation of relative mRNA expression in gill and hemocytes at 24 and 6h of post infection, respectively. These results collectively suggest that AbCyst B is a potent inhibitor of cysteine proteinases and is also potentially involved in immune responses against invading bacterial pathogens in abalone.</description><subject>abalone</subject><subject>Amino Acid Sequence</subject><subject>amino acids</subject><subject>Animals</subject><subject>bacterial artificial chromosomes</subject><subject>Bacterial challenge</subject><subject>bacterial infections</subject><subject>Base Sequence</subject><subject>Cloning, Molecular</subject><subject>complementary DNA</subject><subject>Computer Simulation</subject><subject>Crassostrea gigas</subject><subject>Cystatin B</subject><subject>Cystatin B - chemistry</subject><subject>Cystatin B - genetics</subject><subject>Cystatin B - metabolism</subject><subject>cystatins</subject><subject>Cysteine protease inhibitors</subject><subject>digestive tract</subject><subject>Disk abalone</subject><subject>disulfide bonds</subject><subject>exons</subject><subject>expressed sequence tags</subject><subject>Gastropoda - genetics</subject><subject>Gastropoda - metabolism</subject><subject>Gastropoda - microbiology</subject><subject>gene expression</subject><subject>Gene Expression Regulation</subject><subject>genes</subject><subject>Genome</subject><subject>Haliotis discus</subject><subject>hemocytes</subject><subject>Hemocytes - metabolism</subject><subject>hepatopancreas</subject><subject>Host-Pathogen Interactions</subject><subject>immune response</subject><subject>infection</subject><subject>introns</subject><subject>Models, Molecular</subject><subject>Molecular Sequence Data</subject><subject>muscles</subject><subject>nucleotides</subject><subject>open reading frames</subject><subject>Organ Specificity</subject><subject>papain</subject><subject>Papain - antagonists & inhibitors</subject><subject>Papain - chemistry</subject><subject>Papain inhibition</subject><subject>pathogens</subject><subject>Phylogeny</subject><subject>Proteolysis</subject><subject>screening</subject><subject>Sequence Analysis, DNA</subject><subject>Sequence Homology, Amino Acid</subject><subject>signal peptide</subject><subject>Structural Homology, Protein</subject><subject>testes</subject><subject>Transcriptional analysis</subject><subject>Vibrio parahaemolyticus</subject><subject>Vibrio parahaemolyticus - physiology</subject><issn>0145-305X</issn><issn>1879-0089</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2012</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9Uctu1TAQtRCIXgofwAa8LIuEsfNyxKpU0CJVYgGV2Fl-jFtfkvhiJxXlO_hgHNKyxJvR2Occz5xDyEsGJQPWvt2X1viSA-MltCUweER2THR9ASD6x2QHrG6KCppvR-RZSnvIRzB4So44F52oebMjv89xCqM31NyoqMyM0f9Ssw8TVZOl-PMQMaW1PcTg_ICJLofc6Q2qBuonh-YvITiq6BRucaDmLs1ZZaLv6U0YwxCuF6QuhpFan75TpdUQJqQnF2rwYfZpvTbLQ3nznDxxakj44r4ek6uPH76eXRSXn88_nZ1eFqYWMBeib9HaphOa9do13PbIrKh73jaut7rKb9g0oDulORjVsqrtDLemdp0S2nTVMTnZdPNyPxZMsxzzADgMasKwJMk4h6rlAliGsg1qYkgpopOH6EcV7yQDuYYh9zKHIdcwJLQyh5E5r-7lFz2i_cd4cD8DXm8Ap4JU19EnefUlK7Q5qLxGs377bkNgtuHWY5TJeJwMWh-z7dIG_58B_gCYDaa3</recordid><startdate>20121201</startdate><enddate>20121201</enddate><creator>Premachandra, H.K.A.</creator><creator>Wan, Qiang</creator><creator>Elvitigala, Don Anushka Sandaruwan</creator><creator>De Zoysa, Mahanama</creator><creator>Choi, Cheol Young</creator><creator>Whang, Ilson</creator><creator>Lee, Jehee</creator><general>Elsevier Ltd</general><scope>FBQ</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>20121201</creationdate><title>Genomic characterization and expression profiles upon bacterial infection of a novel cystatin B homologue from disk abalone (Haliotis discus discus)</title><author>Premachandra, H.K.A. ; Wan, Qiang ; Elvitigala, Don Anushka Sandaruwan ; De Zoysa, Mahanama ; Choi, Cheol Young ; Whang, Ilson ; Lee, Jehee</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c480t-896edd578b19bf52d9e1d849265f9db3dd5e550b7ab20ca61367c2dc4f7a8bc73</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2012</creationdate><topic>abalone</topic><topic>Amino Acid Sequence</topic><topic>amino acids</topic><topic>Animals</topic><topic>bacterial artificial chromosomes</topic><topic>Bacterial challenge</topic><topic>bacterial infections</topic><topic>Base Sequence</topic><topic>Cloning, Molecular</topic><topic>complementary DNA</topic><topic>Computer Simulation</topic><topic>Crassostrea gigas</topic><topic>Cystatin B</topic><topic>Cystatin B - chemistry</topic><topic>Cystatin B - genetics</topic><topic>Cystatin B - metabolism</topic><topic>cystatins</topic><topic>Cysteine protease inhibitors</topic><topic>digestive tract</topic><topic>Disk abalone</topic><topic>disulfide bonds</topic><topic>exons</topic><topic>expressed sequence tags</topic><topic>Gastropoda - genetics</topic><topic>Gastropoda - metabolism</topic><topic>Gastropoda - microbiology</topic><topic>gene expression</topic><topic>Gene Expression Regulation</topic><topic>genes</topic><topic>Genome</topic><topic>Haliotis discus</topic><topic>hemocytes</topic><topic>Hemocytes - metabolism</topic><topic>hepatopancreas</topic><topic>Host-Pathogen Interactions</topic><topic>immune response</topic><topic>infection</topic><topic>introns</topic><topic>Models, Molecular</topic><topic>Molecular Sequence Data</topic><topic>muscles</topic><topic>nucleotides</topic><topic>open reading frames</topic><topic>Organ Specificity</topic><topic>papain</topic><topic>Papain - antagonists & inhibitors</topic><topic>Papain - chemistry</topic><topic>Papain inhibition</topic><topic>pathogens</topic><topic>Phylogeny</topic><topic>Proteolysis</topic><topic>screening</topic><topic>Sequence Analysis, DNA</topic><topic>Sequence Homology, Amino Acid</topic><topic>signal peptide</topic><topic>Structural Homology, Protein</topic><topic>testes</topic><topic>Transcriptional analysis</topic><topic>Vibrio parahaemolyticus</topic><topic>Vibrio parahaemolyticus - physiology</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Premachandra, H.K.A.</creatorcontrib><creatorcontrib>Wan, Qiang</creatorcontrib><creatorcontrib>Elvitigala, Don Anushka Sandaruwan</creatorcontrib><creatorcontrib>De Zoysa, Mahanama</creatorcontrib><creatorcontrib>Choi, Cheol Young</creatorcontrib><creatorcontrib>Whang, Ilson</creatorcontrib><creatorcontrib>Lee, Jehee</creatorcontrib><collection>AGRIS</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Developmental and comparative immunology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Premachandra, H.K.A.</au><au>Wan, Qiang</au><au>Elvitigala, Don Anushka Sandaruwan</au><au>De Zoysa, Mahanama</au><au>Choi, Cheol Young</au><au>Whang, Ilson</au><au>Lee, Jehee</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Genomic characterization and expression profiles upon bacterial infection of a novel cystatin B homologue from disk abalone (Haliotis discus discus)</atitle><jtitle>Developmental and comparative immunology</jtitle><addtitle>Dev Comp Immunol</addtitle><date>2012-12-01</date><risdate>2012</risdate><volume>38</volume><issue>4</issue><spage>495</spage><epage>504</epage><pages>495-504</pages><issn>0145-305X</issn><eissn>1879-0089</eissn><abstract>► Complete genomic sequence of cystatin B from disk abalone was identified. ► AbCyt B mRNA was highly expressed in hemocytes, gills, mantle, and digestive tract. ► Bacterial infection caused significant upregulation of AbCyt B expression in gills. ► Recombinant AbCyt B showed strong in vitro papain inhibitory activity.
Cystatins are a large family of cysteine proteinase inhibitors which are involved in diverse biological and pathological processes. In the present study, we identified a gene related to cystatin superfamily, AbCyt B, from disk abalone Haliotis discus discus by expressed sequence tag (EST) analysis and BAC library screening. The complete cDNA sequence of AbCyt B is comprised of 1967 nucleotides with a 306bp open reading frame (ORF) encoding for 101 amino acids. The amino acid sequence consists of a single cystatin-like domain, which has a cysteine proteinase inhibitor signature, a conserved Gly in N-terminal region, QVVAG motif and a variant of PW motif. No signal peptide, disulfide bonds or carbohydrate side chains were identified. Analysis of deduced amino acid sequence revealed that AbCyt B shares up to 44.7% identity and 65.7% similarity with the cystatin B genes from other organisms. The genomic sequence of AbCyt B is approximately 8.4Kb, consisting of three exons and two introns. Phylogenetic tree analysis showed that AbCyt B was closely related to the cystatin B from pacific oyster (Crassostrea gigas) under the family 1.Functional analysis of recombinant AbCyt B protein exhibited inhibitory activity against the papain, with almost 84% inhibition at a concentration of 3.5μmol/L. In tissue expression analysis, AbCyt B transcripts were expressed abundantly in the hemocyte, gill, mantle, and digestive tract, while weakly in muscle, testis, and hepatopancreas. After the immune challenge with Vibrio parahemolyticus, the AbCyt B showed significant (P<0.05) up-regulation of relative mRNA expression in gill and hemocytes at 24 and 6h of post infection, respectively. These results collectively suggest that AbCyst B is a potent inhibitor of cysteine proteinases and is also potentially involved in immune responses against invading bacterial pathogens in abalone.</abstract><cop>United States</cop><pub>Elsevier Ltd</pub><pmid>22878425</pmid><doi>10.1016/j.dci.2012.06.010</doi><tpages>10</tpages></addata></record> |
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| subjects | abalone Amino Acid Sequence amino acids Animals bacterial artificial chromosomes Bacterial challenge bacterial infections Base Sequence Cloning, Molecular complementary DNA Computer Simulation Crassostrea gigas Cystatin B Cystatin B - chemistry Cystatin B - genetics Cystatin B - metabolism cystatins Cysteine protease inhibitors digestive tract Disk abalone disulfide bonds exons expressed sequence tags Gastropoda - genetics Gastropoda - metabolism Gastropoda - microbiology gene expression Gene Expression Regulation genes Genome Haliotis discus hemocytes Hemocytes - metabolism hepatopancreas Host-Pathogen Interactions immune response infection introns Models, Molecular Molecular Sequence Data muscles nucleotides open reading frames Organ Specificity papain Papain - antagonists & inhibitors Papain - chemistry Papain inhibition pathogens Phylogeny Proteolysis screening Sequence Analysis, DNA Sequence Homology, Amino Acid signal peptide Structural Homology, Protein testes Transcriptional analysis Vibrio parahaemolyticus Vibrio parahaemolyticus - physiology |
| title | Genomic characterization and expression profiles upon bacterial infection of a novel cystatin B homologue from disk abalone (Haliotis discus discus) |
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