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Channel catfish (Ictalurus punctatus Rafinesque, 1818) tetraspanin membrane protein family: Identification, characterization and expression analysis of CD63 cDNA
Veterinary Immunology and Immunopathology 133 (2010) 302–308
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Technical report
Channel catfish (Ictalurus punctatus Rafinesque, 1818) tetraspanin membrane protein family: Identification, characterization and expression analysis of CD63 cDNA Hung-Yueh Yeh *, Phillip H. Klesius United States Department of Agriculture, Agricultural Research Service, Aquatic Animal Health Research Unit, 990 Wire Road, Auburn, AL 36832-4352, United States
A R T I C L E I N F O
A B S T R A C T
Article history: Received 5 June 2009 Received in revised form 23 July 2009 Accepted 3 August 2009
CD63, known as lysosome associated membrane protein 3 (LAMP-3), is a member of the tetraspanin integral membrane protein family. This protein plays many important roles in immuno-physiological functions. In this communication, we report the identification, characterization, and expression analysis of the channel catfish CD63 transcript. The complete nucleic acid sequence of channel catfish CD63 cDNA was comprised of 1159 nucleotides, including an open reading frame, which appears to encode a putative peptide of 237-amino-acid residues. Like other tetraspanin proteins, the channel catfish CD63 peptide can be divided into domains, including four transmembrane domains, three intracellular domains, and one of each small and large extracellular loops. The channel catfish CD63 peptide shares 52–55% identity among fish counterparts, but only 43–46% identity among mammalian counterparts. The characteristic Cys-Cys-Gly motif and four Cys residues in the large extracellular loop were conserved. The channel catfish CD63 transcript was detected by RT-PCR in spleen, anterior kidney, liver, intestine, skin and gill. This result provides important information for further elucidating CD63 functions in channel catfish. Published by Elsevier B.V.
Keywords: CD63 Tetraspanin protein Lysosome associated membrane protein 3 LAMP-3 Channel catfish Ictalurus punctatus
The tetraspanin protein superfamily, which consists of more than 30 protein members in humans, can be divided into four sub-families: CD-non63, CD-63, uroplakin and RDS, and plays many important roles in cell–cell and matrix–cell interactions (Hemler, 2005; Levy and Shoham, 2005; Huang et al., 2005; Berditchevski and Odintsova, 2007; Pols and Klumperman, 2009). We are interested in CD63, because we observed that the CD63 transcript was up-regulated during the early stages of Edwardsiella ictaluri infection in channel catfish ovary cells (ATCC CRL-2772) (Yeh and Klesius, unpublished data). CD63, originally known as platelet glycoprotein 40 or melanoma antigen 491 (Hotta et al., 1988; Metzelaar et al., 1989, 1991), is a lysosomal membrane glycoprotein
* Corresponding author. Tel.: +1 334 887 3741; fax: +1 334 887 2983. E-mail address: [email protected] (H.-Y. Yeh). 0165-2427/$ – see front matter . Published by Elsevier B.V. doi:10.1016/j.vetimm.2009.08.006
(Nieuwenhuis et al., 1987; Metzelaar et al., 1991; Schro¨der et al., 2009). This molecule is found as granules in many types of blood cells and endo-/epithelial cells such as dense granules in platelets, a-granules in megakaryocytes, cytotoxic T-cell granules in T cells, azurophil granules in neutrophils, Weibel–Palade bodies in vascular endothelial cells, and eosinophil secretory granules in B cells, dendritic cells and epithelial cells (Nieuwenhuis et al., 1987; Peters et al., 1991a,b; Nishibori et al., 1993; Escola et al., 1996, 1998; Heijnen et al., 1998; Kobayashi et al., 2000, 2002; van der Wel et al., 2003). Upon cell activation, CD63 is mobilized to the cell surface where it is involved in many physiological processes (Berditchevski and Odintsova, 2007; Pols and Klumperman, 2009; Schro¨der et al., 2009). For example, CD63 interacts with MHC class II-peptide complexes in dendritic endosomes, and then cells undergo development and maturation where the CD63–MHCII complexes are
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redistributed to the cell surface for presentation to T cells (Engering and Pieters, 2001; Engering et al., 2003). On the other hand, CD63 plays roles in establishment of intracellular pathogen infection. Beatty (2006, 2008) has demonstrated that trafficking of CD63 multivesicular bodies to the chlamydial inclusions is required for providing the essential nutrients from host cells for Chlamydia development in inclusions. Another example is that CD63 is critical in supporting HIV replication in macrophages besides well-studied cellular factors CD4 and CXCR4 (Chen et al., 2008). Our interest in gene expression in channel catfish (Ictalurus punctatus Rafinesque, 1818) after Edwardsiella ictaluri infection prompted us to identify and characterize tetraspanin cDNAs. We had previously characterized channel catfish CD81 cDNA (Yeh and Klesius, 2009b). In this communication, we report the identification, characterization and expression analysis of channel catfish CD63 cDNA. Channel catfish (NWAC 103 strain) were used in this study according to the Guidelines for the Use of Fish in Research (Nickum et al., 2004). The protocol for experimental use of catfish was approved by the Institutional Animal Care and Use Committee, Aquatic Animal Health Research Unit, Agricultural Research Service, U.S. Department of Agriculture, Auburn, AL. Total RNA from channel catfish tissues was purified by using a Tri reagent (Molecular Research Center, Inc., Cincinnati, OH) per the manufacturer’s instruction. The quality and quantity of total RNA were determined with an Agilent Bioanalyzer using RNA 1200 chips (Agilent Technologies, Santa Clara, CA). Both 50 - and 30 -RACE libraries were constructed by using a GeneRacer kit (Invitrogen, Carlsbad, CA) according to the protocol provided in the kit. Oligonucleotide primers for PCR amplification are listed in Table 1. The amplified PCR fragments were purified and cloned into a pSC vector (Agilent Technologies) per the manufacturer’s instruction. At least six colonies per product were randomly selected and cultured in Wu medium (www.plantgenomics.iastate.edu/protocols/plasmid_isolation.pdf) in 96-well plates for sequencing according to the standard procedures (Sambrook et al., 1989). DNA sequencing on both strands was carried out at the USDA ARS Genomics and Bioinformatics Research Unit (Stoneville, MS) as described previously (Yeh and Klesius, 2007a,b, 2009c). The amino acid sequence was deduced by using the Transeq program (Rice et al., 2000), and aligned
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with other CD63 amino acid sequences deposited in GenBank using ClustalW2 software (Larkin et al., 2007). ExPASy server (Gasteiger et al., 2005) was used to calculate the CD63 peptide molecular mass and pI. Transmembrane topology and signal peptide of the CD63 peptide were predicted via Phobius Web server (Ka¨ll et al., 2007) Phylogenetic relationships of CD63 amino acid sequences from various species were analyzed by the MEGA 4.0 software (Tamura et al., 2007) based on the ClustalW2 alignment results. RT-PCR assays for detection of CD63 in channel catfish tissues were carried out by a two-step procedure described previously (Yeh and Klesius, 2007b, 2008a,b). The transcripts were amplified up to the log phase of PCR amplification. The b-actin transcript was used as an internal control. Images were documented by a Kodak Gel Logic 440 Imaging System (Eastman Kodak Co., Rochester, NY), and processed using the ImageJ software (Abramoff et al., 2004). In our preliminary study, we partially identified the channel catfish CD63 expressed sequence tag (EST) by subtractive suppression hybridization (Yeh and Klesius, unpublished data). Based on this EST, we designed the CD63-specific primers in conjugation with the GeneRacer primers (Table 1) to PCR amplify both 50 - and 30 -end cDNA (Frohman et al., 1988). The full-length CD63 cDNA consists of 1159 nucleotides, including 50 - and 30 untranslated regions (UTR) and an open reading frame (GenBank accession no. FJ899742). A Kozak sequence (Kozak, 1987) was found in 50 -UTR, while three mRNA canonical features were observed in the 30 -UTR. A 714nucleotide open reading frame appears to encode a 237amino-acid peptide with a calculated molecular mass of 25.9 kDa and pI of 7.76 at pH 7.0. The deduced channel catfish CD63 had four potential N-glycosylation sites at Asn123, Asn127, Asn148 and Asn169 in the large extracellular loop (Fig. 1). When we compared the deduced channel catfish CD63 peptide with those from other species deposited in GenBank, we observed that, like the mammalian counterparts (Hotta et al., 1988; Metzelaar et al., 1991), channel catfish CD63 is a transmembrane protein, which can be structurally divided into four transmembrane regions, three cytoplasmic domains and two (small and large) extracellular loops (Fig. 1). Unlike mammalian CD63, which share more than 77% homology, the deduced channel catfish CD63 shares 52–55% identity among fish CD63 and 43–46% identity among mammalian counter-
Table 1 Oligonucleotide primers used for PCR amplification in this study. Primer CD63-F CD63-R CD63-222R CD63-497R GeneRacer 50 Primer (Invitrogen) GeneRacer 30 Primer (Invitrogen) b-Actin-F b-Actin-R
Sequence 0
0
5 -TGGAGGGTGGAATGAAGTGC-3 50 -GATCCGGTGTTGGCTAGCTC-30 50 -CTTTCCACGCTCCACAACAGCCGAAG-30 50 -GCAGCAGGAGTCGGGAACGGAGTTCT-30 50 -CGACTGGAGCACGAGGACACTGA-30 50 -GCTGTCAACGATACGCTACGTAACG-30 50 -GACTTCGAGCAGGAGATGGG-30 50 -AACCTCTCATTGCCAATGGTG-30
Direction
Tm (8C)
Forward Reverse Reverse Reverse Forward Reverse Forward Reverse
67 66 74 74 74 78 72 69
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Fig. 2. Comparison of the channel catfish large extracellular loops from the tetraspanin members CD63 and CD81. The conserved Cys residues and Cys-CysGly motif are in boldface. GenBank accession numbers are as follows: CD63, FJ899742 and CD81, FJ205473 (Yeh and Klesius, 2009b).
Table 2 Identity of CD63 amino acid sequences among species (%)a.
Atlantic salmon Rainbow trout Spotted green pufferfish Zebrafish Western clawed frog Mouse Rat Cattle Horse Rabbit Cat Human a
Channel catfish
Atlantic salmon
Rainbow trout
55 55 53 52 52 45 46 46 43 45 43 45
87 67 60 54 44 45 43 43 44 44 45
64 59 51 44 44 43 43 44 44 45
Spotted green pufferfish
59 54 44 45 46 47 46 46 49
Zebrafish
50 43 43 45 43 43 43 45
Western clawed frog
51 52 54 52 52 50 53
Mouse
Rat
Cattle
Horse
Rabbit
Cat
96 78 78 79 78 79
77 77 79 77 78
81 86 83 85
85 83 84
86 87
89
Identity in percentage between two species was calculated by the ClustalW2 program (Larkin et al., 2007) via http://www.ebi.ac.uk.
parts (Table 2). As shown in Fig. 1, we found that although the low degree of conservation between fish and mammalian CD63, the following CD63 characteristics are conserved (amino acid numbering after channel catfish): (1) the Cys143-Cys144-Gly145 motif in the large extracellular loop (LEL), (2) six Cys residues at positions 143, 144, 166, 167, 174 and 190 in the LEL, indicating that the tertiary structure of CD63 may be conserved, (3) the Val162-Pro163-Asp164-Ser165-Cys166-Cys167 motif in the LEL, and (4) the tyrosine-based lysosomal targeting motif (Gly233-Tyr234-Glu235-Val236-Met237) at the carboxyl terminus (Berditchevski, 2001; Berditchevski and Odintsova, 2007). In our previous study, we have identified and characterized channel catfish CD81, another tetraspanin protein member (Yeh and Klesius, 2009b). We compared the relatedness of these two known channel catfish tetraspanin proteins. Overall, they show a relatively low degree of homology over the entire sequences, but show a high degree of conservation in hydrophobic transmembrane domains. In addition, the canonical features-four Cys residues and a Cys-Cys-Gly
motif-in both CD63 and CD81 large extracellular loops are conserved (Fig. 2, boldface), indicating that the disulfide bonds form within the LEL, and thus tertiary structure is conserved throughout evolution (Seigneuret et al., 2001). Like another tetraspanin member CD81 (Yeh and Klesius, 2009b), mammalian CD63 formed a cluster, distinguishable from the fish counterparts (Fig. 3). As expected, the frog is placed between mammals and teleost fish. In addition, due to heterogeneity of fish (Helfman, 2007), fish CD63 formed a loose clade, and channel catfish CD63 fell in the teleost cluster. This finding is in agreement with our previous studies in other channel catfish genes (Yeh and Klesius, 2007b, 2008a,b,c, 2009a). To determine the expression profile of the CD63 transcript in channel catfish tissues, a two-step RT-PCR assay was performed. The PCR amplified products of CD63 and b-actin had 497 and 203 nucleotides, respectively. As seen in Fig. 4, the CD63 transcript was detected in all tissues of fish examined, suggesting that the channel catfish CD63 transcript is constitutively expressed.
Fig. 1. The deduced CD63 amino acid sequence of channel catfish aligned with those of other species’ CD63 deposited in the GenBank database. To maximize the sequence homology, gaps were introduced in the sequences, indicated by (–). Identical amino acids among all species are denoted by (*) below the sequences. The structural domains of CD63 peptides are indicated above the sequences. The potential N-glycosylation sites are underlined. The tyrosinebased lysosomal targeting motif at the carboxyl terminus is boxed. Species and the corresponding GenBank accession numbers are as follows: Atlantic salmon, NP_001134074; cat, NP_001009855; cattle, Q9XSK2; channel catfish, FJ899742; horse, XP_001504828; human, NP_001771; mouse, NP_031679; rabbit, NP_001075668; rainbow trout, NP_001117968; rat, NP_058821; spotted green pufferfish, CAG03950; western clawed frog, NP_001016413; zebrafish, NP_955837.
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Fig. 3. Phylogenetic relationships of CD63 amino acids among 13 species. The sequences from Fig. 1 were used for the tree generation by bootstrap analysis (1000 replications with its value >50%) of the neighbor-joining method conducted in MEGA4 (Tamura et al., 2007). The scale indicates amino acid substitutions.
Further, the ratios of CD63 and b-actin expression in the agarose gel (Fig. 4A) were analyzed by densitometric software associated with the Kodak Gel Logic 440 Imaging System. As seen in Fig. 4B, the different levels of expression of the CD63 transcript among channel catfish tissues were observed. The high level of CD63 expression was found in intestine and anterior kidney, while the lower levels of expression were found in spleen, liver, skin and gill. This result is in agreement with studies that the CD63 transcript is expressed ubiquitously in human tissues (Hotta et al., 1988; Metzelaar et al., 1991; Nieuwenhuis et al., 1987). In conclusion, the channel catfish CD63 transcript was identified, sequenced and characterized. The transcript was detected in all tissues examined. This result provides us with a critical information for further investigation its role in bacterial infection. Experiments for the CD63 expression in an expression system and production of antibodies are under development.
Fig. 4. Expression of the CD63 transcript in channel catfish tissues (n = 4). (A) The sizes of PCR amplified products for CD63 and b-actin were 497 and 203 base pairs, respectively. Spleen, lanes A, G, M and S; anterior kidney, lanes B, H, N and T; liver, lanes C, I, O and U; intestine, lanes D, J, P and V; skin, lanes E, K, Q and W; gill, lanes F, L, R and X. Lane Y, no template control, and lane Z, 100 bp GeneRuler MW ladders (*, 500 bp and **, 1000 bp; Fermentas Life Sciences, Glen Burnie, MD). (B) Ratios of CD63 and b-actin expression in the agarose gel were analyzed with densitometric software. The y-axis represents the relative expression values of CD63 in an arbitrary unit (average of four fish). The x-axis indicates the channel catfish tissues.
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Acknowledgments We are grateful to Mrs. Dorothy B. Moseley of the USDA ARS Aquatic Animal Health Research Unit in Auburn, AL for excellent technical support, and Dr. Brian E. Scheffler and his Bioinformatics Group at the USDA ARS MidSouth Genomics Laboratory in Stoneville, MS for DNA sequencing and bioinformatics. We also thank Dr. Thomas L. Welker (Aquatic Animal Health Research Unit, Auburn, AL) and three anonymous reviewers for critical comments on this manuscript. This study was supported by the USDA Agricultural Research Service CRIS project no. 6420-32000-020-00D. Mention of trade names or commercial products in this paper is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture. References Abramoff, M.D., Magelhaes, P.J., Ram, S.J., 2004. Images processing with ImageJ. Biophotonics Int. 11, 36–42. Beatty, W.L., 2006. Trafficking from CD63-positive late endocytic multivesicular bodies is essential for intracellular development of Chlamydia trachomatis. J. Cell Sci. 119, 350–359. Beatty, W.L., 2008. Late endocytic multivesicular bodies intersect the chlamydial inclusion in the absence of CD63. Infect. Immun. 76, 2872–2881. Berditchevski, F., 2001. Complexes of tetraspanins with integrins: more than meets the eye. J. Cell Sci. 114, 4143–4151. Berditchevski, F., Odintsova, E., 2007. Tetraspanins as regulators of protein trafficking. Traffic 8, 89–96. Chen, H., Dziuba, N., Friedrich, B., von Lindern, J., Murray, J.L., Rojo, D.R., Hodge, T.W., O’Brien, W.A., Ferguson, M.R., 2008. A critical role for CD63 in HIV replication and infection of macrophages and cell lines. Virology 379, 191–196. Engering, A., Pieters, J., 2001. Association of distinct tetraspanins with MHC class II molecules at different subcellular locations in human immature dendritic cells. Int. Immunol. 13, 127–134. Engering, A., Kuhn, L., Fluitsma, D., Hoefsmit, E., Pieters, J., 2003. Differential post-translational modification of CD63 molecules during maturation of human dendritic cells. Eur. J. Biochem. 270, 2412–2420. Escola, J.M., Deleuil, F., Stang, E., Boretto, J., Chavrier, P., Gorvel, J.P., 1996. Characterization of a lysozyme-major histocompatibility complex class II molecule-loading compartment as a specialized recycling endosome in murine B lymphocytes. J. Biol. Chem. 271, 27360–27365. Escola, J.M., Kleijmeer, M.J., Stoorvogel, W., Griffith, J.M., Yoshie, O., Geuze, H.J., 1998. Selective enrichment of tetraspan proteins on the internal vesicles of multivesicular endosomes and on exosomes secreted by human B-lymphocytes. J. Biol. Chem. 273, 20121–20127. Frohman, M.A., Dush, M.K., Martin, G.R., 1988. Rapid production of fulllength cDNAs from rare transcripts: amplification using a single genespecific oligonucleotide primer. Proc. Natl. Acad. Sci. U.S.A. 85, 8998– 9002. Gasteiger, E., Hoogland, C., Gattiker, A., Duvaud, S., Wilkins, M.R., Appel, R.D., Bairoch, A., 2005. Protein identification and analysis tools on the ExPASy server. In: Walker, J.M. (Ed.), The Proteomics Protocols Handbook. Humana Press, pp. 571–607. Helfman, G.S., 2007. Fish Conservation. Island Press, Washington, DC, pp. 3–16. Heijnen, H.F., Debili, N., Vainchencker, W., Breton-Gorius, J., Geuze, H.J., Sixma, J.J., 1998. Multivesicular bodies are an intermediate stage in the formation of platelet a-granules. Blood 91, 2313–2325. Hemler, M.E., 2005. Tetraspanin functions and associated microdomains. Nat. Rev. Mol. Cell Biol. 6, 801–811. Hotta, H., Ross, A.H., Huebner, K., Isobe, M., Wendeborn, S., Chao, M.V., Ricciardi, R.P., Tsujimoto, Y., Croce, C.M., Koprowski, H., 1988. Molecular cloning and characterization of an antigen associated with early stages of melanoma tumor progression. Cancer Res. 48, 2955–2962. Huang, S., Yuan, S., Dong, M., Su, J., Yu, C., Shen, Y., Xie, X., Yu, Y., Yu, X., Chen, S., Zhang, S., Pontarotti, P., Xu, A., 2005. The phylogenetic analysis of tetraspanins projects the evolution of cell–cell interactions from unicellular to multicellular organisms. Genomics 86, 674–684.
307
Ka¨ll, L., Krogh, A., Sonnhammer, E.L.L., 2007. Advantages of combined transmembrane topology and signal peptide prediction—the Phobius web server. Nucl. Acid Res. 35, W429–W432. Kobayashi, T., Vischer, U.M., Rosnoblet, C., Lebrand, C., Lindsay, M., Parton, R.G., Kruithof, E.K., Gruenberg, J., 2000. The tetraspanin CD63/lamp3 cycles between endocytic and secretory compartments in human endothelial cells. Mol. Biol. Cell 11, 1829–1843. Kobayashi, T., Beuchat, M.H., Chevallier, J., Makino, A., Mayran, N., Escola, J.M., Lebrand, C., Cosson, P., Kobayashi, T., Gruenberg, J., 2002. Separation and characterization of late endosomal membrane domains. J. Biol. Chem. 277, 32157–32164. Kozak, M., 1987. An analysis of 50 -noncoding sequences from 699 vertebrate messenger RNAs. Nucl. Acids Res. 15, 8125–8148. Larkin, M.A., Blackshields, G., Brown, N.P., Chenna, R., McGettigan, P.A., McWilliam, H., Valentin, F., Wallace, I.M., Wilm, A., Lopez, R., Thompson, J.D., Gibson, T.J., Higgins, D.G., 2007. ClustalW and ClustalX version 2. Bioinformatics 23, 2947–2948. Levy, S., Shoham, T., 2005. Protein–protein interactions in the tetraspanin web. Physiology 20, 218–224. Metzelaar, M.J., Sixma, J.J., Nieuwenhuis, H.K., 1989. A new cluster of activation dependent monoclonal antibodies recognizing a 53 kDa lysosome-like granule protein, expressed on the plasma membrane after activation. In: Knapp, W., Dorken, B., Gilks, W.R., Rieber, P., Schmidt, R.E., Stein, H., von dem Borne, A.E.G.K. (Eds.), Leukocyte typing IV: White Cell Differentiation Antigens. Oxford University Press, Oxford, UK, p. 1039. Metzelaar, M.J., Wijngaard, P.L.J., Peters, P.J., Sixma, J.J., Nieuwenhuis, H.K., Clevers, H.C., 1991. CD63 antigen. A novel lysosomal membrane glycoprotein, cloned by a screening procedure for intracellular antigens in eukaryotic cells. J. Biol. Chem. 266, 3239–3245. Nieuwenhuis, H.K., van Oosterhout, J.J., Rozemuller, E., van Iwaarden, F., Sixma, J.J., 1987. Studies with a monoclonal antibody against activated platelets: evidence that a secreted 53,000-molecular weight lysosome-like granule protein is exposed on the surface of activated platelets in the circulation. Blood 70, 838–845. Nickum, J.G., Bart Jr., H.L., Bowser, P.R., Greer, I.E., Hubbs, C., Jenkins, J.A., MacMillan, J.R., Rachlin, J.W., Rose, J.D., Sorensen, P.W., Tomasso, J.R., 2004. Guidelines for the Use of Fishes in Research. American Fisheries Society, Bethesda, MD. Nishibori, M., Cham, B., McNicol, A., Shalev, A., Jain, N., Gerrard, J.M., 1993. The protein CD63 is in platelet dense granules, is deficient in a patient with Hermansky–Pudlak syndrome, and appears identical to granulophysin. J. Clin. Invest. 91, 1775–1782. Peters, P.J., Borst, J., Oorschot, V., Fukuda, M., Kra¨henbu¨hl, O., Tschopp, J., Slot, J.W., Geuze, H.J., 1991a. Cytotoxic T lymphocyte granules are secretory lysosomes, containing both perforin and granzymes. J. Exp. Med. 173, 1099–1109. Peters, P.J., Neefjes, J.J., Oorschot, V., Ploegh, H.L., Geuze, H.J., 1991b. Segregation of MHC class II molecules from MHC class I molecules in the Golgi complex for transport to lysosomal compartments. Nature 349, 669–676. Pols, M.S., Klumperman, J., 2009. Trafficking and function of the tetraspanin CD63. Exp. Cell Res. 315, 1584–1592. Rice, P., Longden, I., Bleasby, A., 2000. EMBOSS: the European molecular biology open software suite. Trends Genet. 16, 276–277. Sambrook, J., Fritsch, E.F., Maniatis, T., 1989. Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY. Schro¨der, J., Lu¨llmann-Rauch, R., Himmerkus, N., Pleines, I., Nieswandt, B., Orinska, Z., Koch-Nolte, F., Schro¨der, B., Bleich, M., Saftig, P., 2009. Deficiency of the tetraspanin CD63 associated with kidney pathology but normal lysosomal function. Mol. Cell. Biol. 29, 1083–1094. Seigneuret, M., Delaguillaumie, A., Lagaudrie`re-Gesbert, C., Conjeaud, H., 2001. Structure of the tetraspanin main extracellular domain. A partially conserved fold with a structurally variable domain insertion. J. Biol. Chem. 276, 40055–40064. Tamura, K., Dudley, J., Nei, M., Kumar, S., 2007. MEGA4: molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol. Biol. Evol. 24, 1596–1599. van der Wel, N.N., Sugita, M., Fluitsma, D.M., Cao, X., Schreibelt, G., Brenner, M.B., Peters, P.J., 2003. CD1 and major histocompatibility complex II molecules follow a different course during dendritic cell maturation. Mol. Biol. Cell 14, 3378–3388. Yeh, H.-Y., Klesius, P.H., 2007a. cDNA cloning, characterization and expression analysis of channel catfish (Ictalurus punctatus, Rafinesque 1818) peroxiredoxin 6 gene. Fish Physiol. Biochem. 33, 233–239. Yeh, H.-Y., Klesius, P.H., 2007b. Molecular cloning and expression of channel catfish, Ictalurus punctatus, complement membrane attack complex inhibitor CD59. Vet. Immunol. Immunopathol. 120, 246– 253.
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Yeh, H.-Y., Klesius, P.H., 2008a. Channel catfish, Ictalurus punctatus, cyclophilin A and B cDNA characterization and expression analysis. Vet. Immunol. Immunopathol. 121, 370–377. Yeh, H.-Y., Klesius, P.H., 2008b. Complete structure, genomic organization, and expression of channel catfish (Ictalurus punctatus, Rafinesque 1818) matrix metalloproteinase-9 gene. Biosci. Biotechnol. Biochem. 72, 702–714. Yeh, H.-Y., Klesius, P.H., 2008c. Molecular cloning, sequencing and characterization of channel catfish (Ictalurus punctatus, Rafinesque 1818) cathepsin S gene. Vet. Immunol. Immunopathol. 126, 382–387.
Yeh, H.-Y., Klesius, P.H., 2009a. Channel catfish, Ictalurus punctatus, cysteine proteinases: cloning, characterisation and expression of cathepsin H and L. Fish Shellfish Immunol. 26, 332–338. Yeh, H.-Y., Klesius, P.H., 2009b. Channel catfish, Ictalurus punctatus Rafinesque 1818, tetraspanin membrane protein family: characterization and expression analysis of CD81 cDNA. Vet. Immunol. Immunopathol. 128, 431–436. Yeh, H.-Y., Klesius, P.H., 2009c. Characterization and tissue expression of channel catfish (Ictalurus punctatus Rafinesque, 1818) ubiquitin carboxyl-terminus hydrolase L5 (UCHL5) cDNA. Mol. Biol. Rep., doi:10.1007/s11033-009-9493-7.
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Technical report
Channel catfish (Ictalurus punctatus Rafinesque, 1818) tetraspanin membrane protein family: Identification, characterization and expression analysis of CD63 cDNA Hung-Yueh Yeh *, Phillip H. Klesius United States Department of Agriculture, Agricultural Research Service, Aquatic Animal Health Research Unit, 990 Wire Road, Auburn, AL 36832-4352, United States
A R T I C L E I N F O
A B S T R A C T
Article history: Received 5 June 2009 Received in revised form 23 July 2009 Accepted 3 August 2009
CD63, known as lysosome associated membrane protein 3 (LAMP-3), is a member of the tetraspanin integral membrane protein family. This protein plays many important roles in immuno-physiological functions. In this communication, we report the identification, characterization, and expression analysis of the channel catfish CD63 transcript. The complete nucleic acid sequence of channel catfish CD63 cDNA was comprised of 1159 nucleotides, including an open reading frame, which appears to encode a putative peptide of 237-amino-acid residues. Like other tetraspanin proteins, the channel catfish CD63 peptide can be divided into domains, including four transmembrane domains, three intracellular domains, and one of each small and large extracellular loops. The channel catfish CD63 peptide shares 52–55% identity among fish counterparts, but only 43–46% identity among mammalian counterparts. The characteristic Cys-Cys-Gly motif and four Cys residues in the large extracellular loop were conserved. The channel catfish CD63 transcript was detected by RT-PCR in spleen, anterior kidney, liver, intestine, skin and gill. This result provides important information for further elucidating CD63 functions in channel catfish. Published by Elsevier B.V.
Keywords: CD63 Tetraspanin protein Lysosome associated membrane protein 3 LAMP-3 Channel catfish Ictalurus punctatus
The tetraspanin protein superfamily, which consists of more than 30 protein members in humans, can be divided into four sub-families: CD-non63, CD-63, uroplakin and RDS, and plays many important roles in cell–cell and matrix–cell interactions (Hemler, 2005; Levy and Shoham, 2005; Huang et al., 2005; Berditchevski and Odintsova, 2007; Pols and Klumperman, 2009). We are interested in CD63, because we observed that the CD63 transcript was up-regulated during the early stages of Edwardsiella ictaluri infection in channel catfish ovary cells (ATCC CRL-2772) (Yeh and Klesius, unpublished data). CD63, originally known as platelet glycoprotein 40 or melanoma antigen 491 (Hotta et al., 1988; Metzelaar et al., 1989, 1991), is a lysosomal membrane glycoprotein
* Corresponding author. Tel.: +1 334 887 3741; fax: +1 334 887 2983. E-mail address: [email protected] (H.-Y. Yeh). 0165-2427/$ – see front matter . Published by Elsevier B.V. doi:10.1016/j.vetimm.2009.08.006
(Nieuwenhuis et al., 1987; Metzelaar et al., 1991; Schro¨der et al., 2009). This molecule is found as granules in many types of blood cells and endo-/epithelial cells such as dense granules in platelets, a-granules in megakaryocytes, cytotoxic T-cell granules in T cells, azurophil granules in neutrophils, Weibel–Palade bodies in vascular endothelial cells, and eosinophil secretory granules in B cells, dendritic cells and epithelial cells (Nieuwenhuis et al., 1987; Peters et al., 1991a,b; Nishibori et al., 1993; Escola et al., 1996, 1998; Heijnen et al., 1998; Kobayashi et al., 2000, 2002; van der Wel et al., 2003). Upon cell activation, CD63 is mobilized to the cell surface where it is involved in many physiological processes (Berditchevski and Odintsova, 2007; Pols and Klumperman, 2009; Schro¨der et al., 2009). For example, CD63 interacts with MHC class II-peptide complexes in dendritic endosomes, and then cells undergo development and maturation where the CD63–MHCII complexes are
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redistributed to the cell surface for presentation to T cells (Engering and Pieters, 2001; Engering et al., 2003). On the other hand, CD63 plays roles in establishment of intracellular pathogen infection. Beatty (2006, 2008) has demonstrated that trafficking of CD63 multivesicular bodies to the chlamydial inclusions is required for providing the essential nutrients from host cells for Chlamydia development in inclusions. Another example is that CD63 is critical in supporting HIV replication in macrophages besides well-studied cellular factors CD4 and CXCR4 (Chen et al., 2008). Our interest in gene expression in channel catfish (Ictalurus punctatus Rafinesque, 1818) after Edwardsiella ictaluri infection prompted us to identify and characterize tetraspanin cDNAs. We had previously characterized channel catfish CD81 cDNA (Yeh and Klesius, 2009b). In this communication, we report the identification, characterization and expression analysis of channel catfish CD63 cDNA. Channel catfish (NWAC 103 strain) were used in this study according to the Guidelines for the Use of Fish in Research (Nickum et al., 2004). The protocol for experimental use of catfish was approved by the Institutional Animal Care and Use Committee, Aquatic Animal Health Research Unit, Agricultural Research Service, U.S. Department of Agriculture, Auburn, AL. Total RNA from channel catfish tissues was purified by using a Tri reagent (Molecular Research Center, Inc., Cincinnati, OH) per the manufacturer’s instruction. The quality and quantity of total RNA were determined with an Agilent Bioanalyzer using RNA 1200 chips (Agilent Technologies, Santa Clara, CA). Both 50 - and 30 -RACE libraries were constructed by using a GeneRacer kit (Invitrogen, Carlsbad, CA) according to the protocol provided in the kit. Oligonucleotide primers for PCR amplification are listed in Table 1. The amplified PCR fragments were purified and cloned into a pSC vector (Agilent Technologies) per the manufacturer’s instruction. At least six colonies per product were randomly selected and cultured in Wu medium (www.plantgenomics.iastate.edu/protocols/plasmid_isolation.pdf) in 96-well plates for sequencing according to the standard procedures (Sambrook et al., 1989). DNA sequencing on both strands was carried out at the USDA ARS Genomics and Bioinformatics Research Unit (Stoneville, MS) as described previously (Yeh and Klesius, 2007a,b, 2009c). The amino acid sequence was deduced by using the Transeq program (Rice et al., 2000), and aligned
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with other CD63 amino acid sequences deposited in GenBank using ClustalW2 software (Larkin et al., 2007). ExPASy server (Gasteiger et al., 2005) was used to calculate the CD63 peptide molecular mass and pI. Transmembrane topology and signal peptide of the CD63 peptide were predicted via Phobius Web server (Ka¨ll et al., 2007) Phylogenetic relationships of CD63 amino acid sequences from various species were analyzed by the MEGA 4.0 software (Tamura et al., 2007) based on the ClustalW2 alignment results. RT-PCR assays for detection of CD63 in channel catfish tissues were carried out by a two-step procedure described previously (Yeh and Klesius, 2007b, 2008a,b). The transcripts were amplified up to the log phase of PCR amplification. The b-actin transcript was used as an internal control. Images were documented by a Kodak Gel Logic 440 Imaging System (Eastman Kodak Co., Rochester, NY), and processed using the ImageJ software (Abramoff et al., 2004). In our preliminary study, we partially identified the channel catfish CD63 expressed sequence tag (EST) by subtractive suppression hybridization (Yeh and Klesius, unpublished data). Based on this EST, we designed the CD63-specific primers in conjugation with the GeneRacer primers (Table 1) to PCR amplify both 50 - and 30 -end cDNA (Frohman et al., 1988). The full-length CD63 cDNA consists of 1159 nucleotides, including 50 - and 30 untranslated regions (UTR) and an open reading frame (GenBank accession no. FJ899742). A Kozak sequence (Kozak, 1987) was found in 50 -UTR, while three mRNA canonical features were observed in the 30 -UTR. A 714nucleotide open reading frame appears to encode a 237amino-acid peptide with a calculated molecular mass of 25.9 kDa and pI of 7.76 at pH 7.0. The deduced channel catfish CD63 had four potential N-glycosylation sites at Asn123, Asn127, Asn148 and Asn169 in the large extracellular loop (Fig. 1). When we compared the deduced channel catfish CD63 peptide with those from other species deposited in GenBank, we observed that, like the mammalian counterparts (Hotta et al., 1988; Metzelaar et al., 1991), channel catfish CD63 is a transmembrane protein, which can be structurally divided into four transmembrane regions, three cytoplasmic domains and two (small and large) extracellular loops (Fig. 1). Unlike mammalian CD63, which share more than 77% homology, the deduced channel catfish CD63 shares 52–55% identity among fish CD63 and 43–46% identity among mammalian counter-
Table 1 Oligonucleotide primers used for PCR amplification in this study. Primer CD63-F CD63-R CD63-222R CD63-497R GeneRacer 50 Primer (Invitrogen) GeneRacer 30 Primer (Invitrogen) b-Actin-F b-Actin-R
Sequence 0
0
5 -TGGAGGGTGGAATGAAGTGC-3 50 -GATCCGGTGTTGGCTAGCTC-30 50 -CTTTCCACGCTCCACAACAGCCGAAG-30 50 -GCAGCAGGAGTCGGGAACGGAGTTCT-30 50 -CGACTGGAGCACGAGGACACTGA-30 50 -GCTGTCAACGATACGCTACGTAACG-30 50 -GACTTCGAGCAGGAGATGGG-30 50 -AACCTCTCATTGCCAATGGTG-30
Direction
Tm (8C)
Forward Reverse Reverse Reverse Forward Reverse Forward Reverse
67 66 74 74 74 78 72 69
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Fig. 2. Comparison of the channel catfish large extracellular loops from the tetraspanin members CD63 and CD81. The conserved Cys residues and Cys-CysGly motif are in boldface. GenBank accession numbers are as follows: CD63, FJ899742 and CD81, FJ205473 (Yeh and Klesius, 2009b).
Table 2 Identity of CD63 amino acid sequences among species (%)a.
Atlantic salmon Rainbow trout Spotted green pufferfish Zebrafish Western clawed frog Mouse Rat Cattle Horse Rabbit Cat Human a
Channel catfish
Atlantic salmon
Rainbow trout
55 55 53 52 52 45 46 46 43 45 43 45
87 67 60 54 44 45 43 43 44 44 45
64 59 51 44 44 43 43 44 44 45
Spotted green pufferfish
59 54 44 45 46 47 46 46 49
Zebrafish
50 43 43 45 43 43 43 45
Western clawed frog
51 52 54 52 52 50 53
Mouse
Rat
Cattle
Horse
Rabbit
Cat
96 78 78 79 78 79
77 77 79 77 78
81 86 83 85
85 83 84
86 87
89
Identity in percentage between two species was calculated by the ClustalW2 program (Larkin et al., 2007) via http://www.ebi.ac.uk.
parts (Table 2). As shown in Fig. 1, we found that although the low degree of conservation between fish and mammalian CD63, the following CD63 characteristics are conserved (amino acid numbering after channel catfish): (1) the Cys143-Cys144-Gly145 motif in the large extracellular loop (LEL), (2) six Cys residues at positions 143, 144, 166, 167, 174 and 190 in the LEL, indicating that the tertiary structure of CD63 may be conserved, (3) the Val162-Pro163-Asp164-Ser165-Cys166-Cys167 motif in the LEL, and (4) the tyrosine-based lysosomal targeting motif (Gly233-Tyr234-Glu235-Val236-Met237) at the carboxyl terminus (Berditchevski, 2001; Berditchevski and Odintsova, 2007). In our previous study, we have identified and characterized channel catfish CD81, another tetraspanin protein member (Yeh and Klesius, 2009b). We compared the relatedness of these two known channel catfish tetraspanin proteins. Overall, they show a relatively low degree of homology over the entire sequences, but show a high degree of conservation in hydrophobic transmembrane domains. In addition, the canonical features-four Cys residues and a Cys-Cys-Gly
motif-in both CD63 and CD81 large extracellular loops are conserved (Fig. 2, boldface), indicating that the disulfide bonds form within the LEL, and thus tertiary structure is conserved throughout evolution (Seigneuret et al., 2001). Like another tetraspanin member CD81 (Yeh and Klesius, 2009b), mammalian CD63 formed a cluster, distinguishable from the fish counterparts (Fig. 3). As expected, the frog is placed between mammals and teleost fish. In addition, due to heterogeneity of fish (Helfman, 2007), fish CD63 formed a loose clade, and channel catfish CD63 fell in the teleost cluster. This finding is in agreement with our previous studies in other channel catfish genes (Yeh and Klesius, 2007b, 2008a,b,c, 2009a). To determine the expression profile of the CD63 transcript in channel catfish tissues, a two-step RT-PCR assay was performed. The PCR amplified products of CD63 and b-actin had 497 and 203 nucleotides, respectively. As seen in Fig. 4, the CD63 transcript was detected in all tissues of fish examined, suggesting that the channel catfish CD63 transcript is constitutively expressed.
Fig. 1. The deduced CD63 amino acid sequence of channel catfish aligned with those of other species’ CD63 deposited in the GenBank database. To maximize the sequence homology, gaps were introduced in the sequences, indicated by (–). Identical amino acids among all species are denoted by (*) below the sequences. The structural domains of CD63 peptides are indicated above the sequences. The potential N-glycosylation sites are underlined. The tyrosinebased lysosomal targeting motif at the carboxyl terminus is boxed. Species and the corresponding GenBank accession numbers are as follows: Atlantic salmon, NP_001134074; cat, NP_001009855; cattle, Q9XSK2; channel catfish, FJ899742; horse, XP_001504828; human, NP_001771; mouse, NP_031679; rabbit, NP_001075668; rainbow trout, NP_001117968; rat, NP_058821; spotted green pufferfish, CAG03950; western clawed frog, NP_001016413; zebrafish, NP_955837.
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Fig. 3. Phylogenetic relationships of CD63 amino acids among 13 species. The sequences from Fig. 1 were used for the tree generation by bootstrap analysis (1000 replications with its value >50%) of the neighbor-joining method conducted in MEGA4 (Tamura et al., 2007). The scale indicates amino acid substitutions.
Further, the ratios of CD63 and b-actin expression in the agarose gel (Fig. 4A) were analyzed by densitometric software associated with the Kodak Gel Logic 440 Imaging System. As seen in Fig. 4B, the different levels of expression of the CD63 transcript among channel catfish tissues were observed. The high level of CD63 expression was found in intestine and anterior kidney, while the lower levels of expression were found in spleen, liver, skin and gill. This result is in agreement with studies that the CD63 transcript is expressed ubiquitously in human tissues (Hotta et al., 1988; Metzelaar et al., 1991; Nieuwenhuis et al., 1987). In conclusion, the channel catfish CD63 transcript was identified, sequenced and characterized. The transcript was detected in all tissues examined. This result provides us with a critical information for further investigation its role in bacterial infection. Experiments for the CD63 expression in an expression system and production of antibodies are under development.
Fig. 4. Expression of the CD63 transcript in channel catfish tissues (n = 4). (A) The sizes of PCR amplified products for CD63 and b-actin were 497 and 203 base pairs, respectively. Spleen, lanes A, G, M and S; anterior kidney, lanes B, H, N and T; liver, lanes C, I, O and U; intestine, lanes D, J, P and V; skin, lanes E, K, Q and W; gill, lanes F, L, R and X. Lane Y, no template control, and lane Z, 100 bp GeneRuler MW ladders (*, 500 bp and **, 1000 bp; Fermentas Life Sciences, Glen Burnie, MD). (B) Ratios of CD63 and b-actin expression in the agarose gel were analyzed with densitometric software. The y-axis represents the relative expression values of CD63 in an arbitrary unit (average of four fish). The x-axis indicates the channel catfish tissues.
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Acknowledgments We are grateful to Mrs. Dorothy B. Moseley of the USDA ARS Aquatic Animal Health Research Unit in Auburn, AL for excellent technical support, and Dr. Brian E. Scheffler and his Bioinformatics Group at the USDA ARS MidSouth Genomics Laboratory in Stoneville, MS for DNA sequencing and bioinformatics. We also thank Dr. Thomas L. Welker (Aquatic Animal Health Research Unit, Auburn, AL) and three anonymous reviewers for critical comments on this manuscript. This study was supported by the USDA Agricultural Research Service CRIS project no. 6420-32000-020-00D. Mention of trade names or commercial products in this paper is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture. References Abramoff, M.D., Magelhaes, P.J., Ram, S.J., 2004. Images processing with ImageJ. Biophotonics Int. 11, 36–42. Beatty, W.L., 2006. Trafficking from CD63-positive late endocytic multivesicular bodies is essential for intracellular development of Chlamydia trachomatis. J. Cell Sci. 119, 350–359. Beatty, W.L., 2008. Late endocytic multivesicular bodies intersect the chlamydial inclusion in the absence of CD63. Infect. Immun. 76, 2872–2881. Berditchevski, F., 2001. Complexes of tetraspanins with integrins: more than meets the eye. J. Cell Sci. 114, 4143–4151. Berditchevski, F., Odintsova, E., 2007. Tetraspanins as regulators of protein trafficking. Traffic 8, 89–96. Chen, H., Dziuba, N., Friedrich, B., von Lindern, J., Murray, J.L., Rojo, D.R., Hodge, T.W., O’Brien, W.A., Ferguson, M.R., 2008. A critical role for CD63 in HIV replication and infection of macrophages and cell lines. Virology 379, 191–196. Engering, A., Pieters, J., 2001. Association of distinct tetraspanins with MHC class II molecules at different subcellular locations in human immature dendritic cells. Int. Immunol. 13, 127–134. Engering, A., Kuhn, L., Fluitsma, D., Hoefsmit, E., Pieters, J., 2003. Differential post-translational modification of CD63 molecules during maturation of human dendritic cells. Eur. J. Biochem. 270, 2412–2420. Escola, J.M., Deleuil, F., Stang, E., Boretto, J., Chavrier, P., Gorvel, J.P., 1996. Characterization of a lysozyme-major histocompatibility complex class II molecule-loading compartment as a specialized recycling endosome in murine B lymphocytes. J. Biol. Chem. 271, 27360–27365. Escola, J.M., Kleijmeer, M.J., Stoorvogel, W., Griffith, J.M., Yoshie, O., Geuze, H.J., 1998. Selective enrichment of tetraspan proteins on the internal vesicles of multivesicular endosomes and on exosomes secreted by human B-lymphocytes. J. Biol. Chem. 273, 20121–20127. Frohman, M.A., Dush, M.K., Martin, G.R., 1988. Rapid production of fulllength cDNAs from rare transcripts: amplification using a single genespecific oligonucleotide primer. Proc. Natl. Acad. Sci. U.S.A. 85, 8998– 9002. Gasteiger, E., Hoogland, C., Gattiker, A., Duvaud, S., Wilkins, M.R., Appel, R.D., Bairoch, A., 2005. Protein identification and analysis tools on the ExPASy server. In: Walker, J.M. (Ed.), The Proteomics Protocols Handbook. Humana Press, pp. 571–607. Helfman, G.S., 2007. Fish Conservation. Island Press, Washington, DC, pp. 3–16. Heijnen, H.F., Debili, N., Vainchencker, W., Breton-Gorius, J., Geuze, H.J., Sixma, J.J., 1998. Multivesicular bodies are an intermediate stage in the formation of platelet a-granules. Blood 91, 2313–2325. Hemler, M.E., 2005. Tetraspanin functions and associated microdomains. Nat. Rev. Mol. Cell Biol. 6, 801–811. Hotta, H., Ross, A.H., Huebner, K., Isobe, M., Wendeborn, S., Chao, M.V., Ricciardi, R.P., Tsujimoto, Y., Croce, C.M., Koprowski, H., 1988. Molecular cloning and characterization of an antigen associated with early stages of melanoma tumor progression. Cancer Res. 48, 2955–2962. Huang, S., Yuan, S., Dong, M., Su, J., Yu, C., Shen, Y., Xie, X., Yu, Y., Yu, X., Chen, S., Zhang, S., Pontarotti, P., Xu, A., 2005. The phylogenetic analysis of tetraspanins projects the evolution of cell–cell interactions from unicellular to multicellular organisms. Genomics 86, 674–684.
307
Ka¨ll, L., Krogh, A., Sonnhammer, E.L.L., 2007. Advantages of combined transmembrane topology and signal peptide prediction—the Phobius web server. Nucl. Acid Res. 35, W429–W432. Kobayashi, T., Vischer, U.M., Rosnoblet, C., Lebrand, C., Lindsay, M., Parton, R.G., Kruithof, E.K., Gruenberg, J., 2000. The tetraspanin CD63/lamp3 cycles between endocytic and secretory compartments in human endothelial cells. Mol. Biol. Cell 11, 1829–1843. Kobayashi, T., Beuchat, M.H., Chevallier, J., Makino, A., Mayran, N., Escola, J.M., Lebrand, C., Cosson, P., Kobayashi, T., Gruenberg, J., 2002. Separation and characterization of late endosomal membrane domains. J. Biol. Chem. 277, 32157–32164. Kozak, M., 1987. An analysis of 50 -noncoding sequences from 699 vertebrate messenger RNAs. Nucl. Acids Res. 15, 8125–8148. Larkin, M.A., Blackshields, G., Brown, N.P., Chenna, R., McGettigan, P.A., McWilliam, H., Valentin, F., Wallace, I.M., Wilm, A., Lopez, R., Thompson, J.D., Gibson, T.J., Higgins, D.G., 2007. ClustalW and ClustalX version 2. Bioinformatics 23, 2947–2948. Levy, S., Shoham, T., 2005. Protein–protein interactions in the tetraspanin web. Physiology 20, 218–224. Metzelaar, M.J., Sixma, J.J., Nieuwenhuis, H.K., 1989. A new cluster of activation dependent monoclonal antibodies recognizing a 53 kDa lysosome-like granule protein, expressed on the plasma membrane after activation. In: Knapp, W., Dorken, B., Gilks, W.R., Rieber, P., Schmidt, R.E., Stein, H., von dem Borne, A.E.G.K. (Eds.), Leukocyte typing IV: White Cell Differentiation Antigens. Oxford University Press, Oxford, UK, p. 1039. Metzelaar, M.J., Wijngaard, P.L.J., Peters, P.J., Sixma, J.J., Nieuwenhuis, H.K., Clevers, H.C., 1991. CD63 antigen. A novel lysosomal membrane glycoprotein, cloned by a screening procedure for intracellular antigens in eukaryotic cells. J. Biol. Chem. 266, 3239–3245. Nieuwenhuis, H.K., van Oosterhout, J.J., Rozemuller, E., van Iwaarden, F., Sixma, J.J., 1987. Studies with a monoclonal antibody against activated platelets: evidence that a secreted 53,000-molecular weight lysosome-like granule protein is exposed on the surface of activated platelets in the circulation. Blood 70, 838–845. Nickum, J.G., Bart Jr., H.L., Bowser, P.R., Greer, I.E., Hubbs, C., Jenkins, J.A., MacMillan, J.R., Rachlin, J.W., Rose, J.D., Sorensen, P.W., Tomasso, J.R., 2004. Guidelines for the Use of Fishes in Research. American Fisheries Society, Bethesda, MD. Nishibori, M., Cham, B., McNicol, A., Shalev, A., Jain, N., Gerrard, J.M., 1993. The protein CD63 is in platelet dense granules, is deficient in a patient with Hermansky–Pudlak syndrome, and appears identical to granulophysin. J. Clin. Invest. 91, 1775–1782. Peters, P.J., Borst, J., Oorschot, V., Fukuda, M., Kra¨henbu¨hl, O., Tschopp, J., Slot, J.W., Geuze, H.J., 1991a. Cytotoxic T lymphocyte granules are secretory lysosomes, containing both perforin and granzymes. J. Exp. Med. 173, 1099–1109. Peters, P.J., Neefjes, J.J., Oorschot, V., Ploegh, H.L., Geuze, H.J., 1991b. Segregation of MHC class II molecules from MHC class I molecules in the Golgi complex for transport to lysosomal compartments. Nature 349, 669–676. Pols, M.S., Klumperman, J., 2009. Trafficking and function of the tetraspanin CD63. Exp. Cell Res. 315, 1584–1592. Rice, P., Longden, I., Bleasby, A., 2000. EMBOSS: the European molecular biology open software suite. Trends Genet. 16, 276–277. Sambrook, J., Fritsch, E.F., Maniatis, T., 1989. Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY. Schro¨der, J., Lu¨llmann-Rauch, R., Himmerkus, N., Pleines, I., Nieswandt, B., Orinska, Z., Koch-Nolte, F., Schro¨der, B., Bleich, M., Saftig, P., 2009. Deficiency of the tetraspanin CD63 associated with kidney pathology but normal lysosomal function. Mol. Cell. Biol. 29, 1083–1094. Seigneuret, M., Delaguillaumie, A., Lagaudrie`re-Gesbert, C., Conjeaud, H., 2001. Structure of the tetraspanin main extracellular domain. A partially conserved fold with a structurally variable domain insertion. J. Biol. Chem. 276, 40055–40064. Tamura, K., Dudley, J., Nei, M., Kumar, S., 2007. MEGA4: molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol. Biol. Evol. 24, 1596–1599. van der Wel, N.N., Sugita, M., Fluitsma, D.M., Cao, X., Schreibelt, G., Brenner, M.B., Peters, P.J., 2003. CD1 and major histocompatibility complex II molecules follow a different course during dendritic cell maturation. Mol. Biol. Cell 14, 3378–3388. Yeh, H.-Y., Klesius, P.H., 2007a. cDNA cloning, characterization and expression analysis of channel catfish (Ictalurus punctatus, Rafinesque 1818) peroxiredoxin 6 gene. Fish Physiol. Biochem. 33, 233–239. Yeh, H.-Y., Klesius, P.H., 2007b. Molecular cloning and expression of channel catfish, Ictalurus punctatus, complement membrane attack complex inhibitor CD59. Vet. Immunol. Immunopathol. 120, 246– 253.
308
H.-Y. Yeh, P.H. Klesius / Veterinary Immunology and Immunopathology 133 (2010) 302–308
Yeh, H.-Y., Klesius, P.H., 2008a. Channel catfish, Ictalurus punctatus, cyclophilin A and B cDNA characterization and expression analysis. Vet. Immunol. Immunopathol. 121, 370–377. Yeh, H.-Y., Klesius, P.H., 2008b. Complete structure, genomic organization, and expression of channel catfish (Ictalurus punctatus, Rafinesque 1818) matrix metalloproteinase-9 gene. Biosci. Biotechnol. Biochem. 72, 702–714. Yeh, H.-Y., Klesius, P.H., 2008c. Molecular cloning, sequencing and characterization of channel catfish (Ictalurus punctatus, Rafinesque 1818) cathepsin S gene. Vet. Immunol. Immunopathol. 126, 382–387.
Yeh, H.-Y., Klesius, P.H., 2009a. Channel catfish, Ictalurus punctatus, cysteine proteinases: cloning, characterisation and expression of cathepsin H and L. Fish Shellfish Immunol. 26, 332–338. Yeh, H.-Y., Klesius, P.H., 2009b. Channel catfish, Ictalurus punctatus Rafinesque 1818, tetraspanin membrane protein family: characterization and expression analysis of CD81 cDNA. Vet. Immunol. Immunopathol. 128, 431–436. Yeh, H.-Y., Klesius, P.H., 2009c. Characterization and tissue expression of channel catfish (Ictalurus punctatus Rafinesque, 1818) ubiquitin carboxyl-terminus hydrolase L5 (UCHL5) cDNA. Mol. Biol. Rep., doi:10.1007/s11033-009-9493-7.