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United States Department of Agriculture

Agricultural Research Service

Research Project: MOLECULAR ANALYSIS OF VIRULENCE DETERMINANTS OF SELECT BACTERIA IN FISH DISEASES

Location: Aquatic Animal Health Research

Title: Molecular Cloning, Sequencing and Characterization of Channel Catfish (Ictalurus punctatus, Rafinesque 1818) Cathepsin S Gene

Authors
item YEH, HUNG-YUEH
item Klesius, Phillip

Submitted to: Veterinary Immunology and Immunopathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: July 18, 2008
Publication Date: February 17, 2009
Repository URL: http://hdl.handle.net/10113/22025
Citation: Yeh, H., Klesius, P.H. 2008. Molecular cloning, sequencing and characterization of channel catfish (Ictalurus punctatus, Rafinesque 1818) cathepsin S gene. Veterinary Immunology and Immunopathology. 126:382-387.

Interpretive Summary: Channel catfish production is the most important aquacultural industry in the southeastern United States, generating the value of more than 450 million dollars annually. We sequenced, identified and characterized the channel catfish cathepsin S (CTSS). CTSS is a lysosomal cysteine endopeptidase of the papain family. This enzyme digests the invariant chain molecules so that antigenic peptides are able to load on the class II-associated invariant chain peptide of MHC. The complexes can subsequently be presented to the CD4 cell surface. The complete channel catfish CTSS cDNA comprised 1530 bp including a 96-bp 5’-untranslated, a 990-bp open reading frame and a 444-bp 3’-untranslated regions. The open reading frame encodes a putative protein of 329 amino acid residues with calculated molecular mass (without potential glycosylation) of 36.7 kDa and pI of 5.96. The degree of identity of the channel catfish CTSS amino acid sequence to the counterparts of other species ranged from 56.6% to 68.5%. This result will provide fundamental information for further exploration of the role of CTSS in fish pathophysiology as well as for the production of monoclonal/ polyclonal antibodies for channel catfish CTSS.

Technical Abstract: Cathepsin S is a lysosomal cysteine endopeptidase of the papain family. Our preliminary results showed the up-regulation of cathepsin S (CTSS) transcript during the early stage of Edwardsiella ictaluri infection. This prompted us to speculate that the CTSS may play a role in infection. In this report, we identified, sequenced and characterized the channel catfish CTSS cDNA. Total RNA from tissues was isolated and cDNA libraries were constructed by the rapid amplification cDNA end (RACE) method. The gene-specific primers in conjunction with the RACE primers were used to PCR amplify 5’- and 3’-ends of the CTSS transcript. The complete channel catfish CTSS cDNA comprised 1530 nucleotides including a 96-nucleotides 5’-untranslated, a 990-nucleotides open reading frame and a 444-nucleotides 3’-untranslated regions. The open reading frame potentially encoded a protein of 329 amino acid residues with calculated molecular mass of 36.7 kDa and pI of 5.96. The degree of conservation of the channel catfish CTSS amino acid sequence to the counterparts of other species ranged from 56.6% to 68.5%. The result provides important information for further exploring the roles of channel catfish CTSS in antigen processing.

Last Modified: 9/10/2014
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