Skip to main content
SpringerLink
Log in

Identification and characterization of a FasL-like protein and cDNAs encoding the channel catfish death-inducing signaling complex

  • Original Paper
  • Published:
Immunogenetics Aims and scope Submit manuscript

Abstract

To elucidate cytolytic mechanisms in the channel catfish, lysates from catfish lymphoid and fibroblast cell lines were screened by Western blot analysis using a panel of antibodies reactive with components of the mammalian apoptotic pathway. Strong reactivity with three proteins (approximate Mr 70,000, 37,000, and 15,000) was seen using an antibody targeted to mammalian Fas ligand (FasL). The sizes of the two smaller proteins are consistent with their tentative designation as membrane-bound (37,000 Mr) and soluble (15,000 Mr) FasL. Treatments known to induce FasL in mammalian systems (e.g., PMA/calcium ionophore, UV-irradiation) induced expression of the 37,000-Mr protein in catfish T-cell lines. Moreover, expression of the 37,000-Mr protein in clonal T cells was up-regulated by increasing cell density. At the nucleotide level, homologues of Fas receptor (FasR), FADD, and caspase 8 were identified and characterized. These gene products likely constitute the teleost equivalent of the death-inducing signaling complex (DISC). FADD was constitutively expressed in all (T, B, macrophage, and fibroblast) cell lines examined as well as in peripheral blood lymphocytes (PBL), whereas FasR and caspase 8 were expressed in all cell lines except CCO, a FasL-positive fibroblast line. In contrast to FasL, expression of FasR and caspase 8 was inversely proportional to cell density. Collectively these studies identified four membrane-proximal proteins involved in the initiation of apoptosis in channel catfish and suggest that mechanisms of cell-mediated cytotoxicity in teleosts are similar to those used by mammals.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2a-c
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  • Algeciras-Schimnich A, Paya C (2000) Transcriptional regulation of the FasL gene. In: Sitkovsky M, Henkart P (eds) Cytotoxic cells: basic mechanisms and medical applications. Lippincott, Williams & Wilkins, Philadelphia, pp 239–247

    Google Scholar 

  • Altschul S, Maden T, Schaffer A, Zhang J, Zhang Z, Miller W, Lipman D (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    CAS  PubMed  Google Scholar 

  • Ayroldi E, D’Adamio F, Zollo O, Agostini M, Moraca R, Cannarile L, Migliorati G, Delfino D, Riccardi C (1999) Cloning and expression of a short Fas ligand: a new alternatively spliced product of the mouse Fas ligand. Blood 94(10):3456–3467

    CAS  PubMed  Google Scholar 

  • Bajorath J (1999) Identification of the ligand binding site in Fas (CD95) and analysis of Fas–ligand interactions. Proteins: Struct Funct Genet 35:475–482

    Article  CAS  Google Scholar 

  • Banner D, D’Acry A, Janes W, Gentz R, Schoenfeld H-J, Broger C, Loetscher H, Lesslauer W (1993) Crystal structure of the soluble human 55 kD TNF receptor-human TNFβ complex: implications for TNF receptor activation. Cell 73:431–445

    Article  CAS  PubMed  Google Scholar 

  • Bazan J (1993) Emerging families of cytokines and receptors. Curr Biol 3:603–606

    Article  CAS  PubMed  Google Scholar 

  • Bishop G, Taylor S, Jaso-Friedmann L, Evans D (2002) Mechanisms of nonspecific cytotoxic regulation of apoptosis: cytokine-like activity of Fas ligand. Fish Shellfish Immunol 13:47–67

    Article  CAS  PubMed  Google Scholar 

  • Bodmer J-L, Schneider P, Tschopp J (2002) The molecular architecture of the TNF superfamily. Trends Biochem Sci 271:19–26

    Article  Google Scholar 

  • Bouchier-Hayes L, Martin SJ (2002) CARD games in apoptosis and immunity. EMBO Rep 3:616–621

    Article  CAS  PubMed  Google Scholar 

  • Bowser P, Plumb J (1980) Growth rates of a new cell line from the channel catfish ovary and channel catfish virus replication at different temperatures. Can J Fish Aquat Sci 37:871–873

    Google Scholar 

  • Bremner T, Chatterjee D, Han Z, Tsan M, Wyche J (1999) THP-1 monocytic leukemia cells express Fas ligand constitutively and kill Fas-positive Jurkat cells. Leuk Res 23:865–870

    Article  CAS  PubMed  Google Scholar 

  • Caricchio R, Reap E, Cohen P (1998) Fas/Fas ligand interactions are involved in ultraviolet-B-induced human lymphocyte apoptosis. J Immunol 161:241–251

    CAS  PubMed  Google Scholar 

  • Carlson RL, Evans DL, Graves SS (1985) Nonspecific cytotoxic cells in fish (Ictalurus punctatus) V. metabolic requirements of lysis. Dev Comp Immunol 9:271–280

    Article  CAS  PubMed  Google Scholar 

  • Chinnaiyan A, Chaudhary D, O’Rourke K, Koonin E, Dixit V (1997) Role of CED-4 in the activation of CED-3. Nature 388:728–729

    Article  CAS  PubMed  Google Scholar 

  • Cohen G (1997) Caspases: the executioners of apoptosis. J Biochem 326:1–16

    CAS  Google Scholar 

  • Cuesta A, Esteban A, Meseguer J (2003) Identification of a FasL-like molecule in leucocytes of the teleost fish gilthead seabream (Sparus aurata L.). Dev Comp Immunol 27:21–27

    Article  CAS  PubMed  Google Scholar 

  • Darnell J, Lodish H, Baltimore D (1990) Molecular cell biology. Scientific American/Freeman, New York

    Google Scholar 

  • Duan H, Dixit V (1997) RAIDD is a new ‘death’ adaptor protein. Nature 385:86–89

    Article  CAS  PubMed  Google Scholar 

  • Enari M (1998) A caspase-activated DNase that degrades DNA during apoptosis, and its inhibitor ICAD. Nature 391:43–50

    Article  CAS  PubMed  Google Scholar 

  • Evans D, Graves S, Cobb D, Dawe D (1984) Nonspecific cytotoxic cells in fish (Ictalurus punctatus). II. Parameters of target cell lysis and specificity. Dev Comp Immunol 8:303–312

    Article  CAS  PubMed  Google Scholar 

  • Evans D, Taylor S, Leary J, Bishop G, Eldar A, Jaso-Friedmann L (2000) In vivo activation of tilapia nonspecific cytotoxic cells by Streptococcus iniae and amplification with apoptosis regulatory factor(s). Fish Shellfish Immunol 10:419–434

    Article  CAS  PubMed  Google Scholar 

  • Evans D, Leary H, Jaso-Friedmann L (2001) Nonspecific cytotoxic cells and innate immunity: regulation by programmed cell death. Dev Comp Immunol 25:791–805

    Article  CAS  PubMed  Google Scholar 

  • Falquet L, Pagni M, Bucher P, Hulo N, Sigrist CJ, Hofmann K, Bairoch A (2002) The PROSITE database, its status in 2002. Nucleic Acids Res 2002:235–238

    Article  Google Scholar 

  • Graves S, Evans D, Dawe D (1985) Antiprotozoan activity of nonspecific cytotoxic cells (NCC) from the channel catfish (Ictalurus punctatus). J Immunol 134:78–85

    CAS  PubMed  Google Scholar 

  • Hawke N, Yoder J, Litman G (1999) Expanding our understanding of immunoglobulin, T-cell antigen receptor, and novel immune-type receptor genes: a subset of the immunoglobulin gene superfamily. Immunogenetics 50:124–133

    Article  CAS  PubMed  Google Scholar 

  • Hofmann K, Stoffel W (1993) Tmbase: a database of membrane spanning protein segments. Biol Chem Hoppe-Seyler 374:166

    Google Scholar 

  • Hofmann K, Bucher P, Tschopp J (1997) The CARD domain is a new apoptotic signaling motif. Trends Biochem Sci 22:15–156

    Article  PubMed  Google Scholar 

  • Hogan R, Taylor W, Cuchens M, Naftel J, Clem LW, Miller NW, Chinchar VG (1999) Induction of target cell apoptosis by channel catfish cytotoxic cells. Cell Immunol 195:110–118

    Article  CAS  PubMed  Google Scholar 

  • Irmler M, Hofmann K, Vaux D, Tschopp J (1997) Direct physical interactions between the Caenorhabditis elegans ‘death proteins’ CED-3 and CED-4. FEBS Lett 406:189–190

    Article  CAS  PubMed  Google Scholar 

  • Jaso-Friedmann L, Leary J, Evans D (2000) Role of nonspecific cytotoxic cells in the induction of programmed cell death of pathogenic protozoans: participation of the Fas ligand–Fas receptor system. Exp Parasitol 96:75–88

    Article  CAS  PubMed  Google Scholar 

  • Jones E, Stuart D, Walker N (1989) Structure of tumor necrosis factor. Nature 338:225–228

    Article  CAS  PubMed  Google Scholar 

  • Kiener P, Davis P, Rankin B, Klebanoff S, Ledbetter J, Starling G, Liles WC (1997) Human monocytic cells contain high levels of intracellular Fas ligand. J Immunol 159:1594–1598

    CAS  PubMed  Google Scholar 

  • Krammer P, Debatin K-M (1992) A gene for a cell surface molecule known as Fas or APO-1 that can trigger apoptosis has been identified as the target of mutations that cause lymphoproliferation and autoimmunity in mice. Curr Biol 2:383–385

    Article  CAS  PubMed  Google Scholar 

  • Kuhn K, Hashimoto S, Lotz M (1999) Cell density modulates apoptosis in human articular chondrocytes. J Cell Physiol 180:439–447

    Article  CAS  PubMed  Google Scholar 

  • Kumar S, Hedges S (1998) A molecular timescale for vertebrate evolution. Nature 392:917–920

    Article  CAS  PubMed  Google Scholar 

  • Kumar S, Tamura K, Jakobsen I, Nei M (2001) MEGA2: molecular evolutionary genetics analysis software. Bioinformatics 17:1244–1245

    Article  CAS  PubMed  Google Scholar 

  • Laemmli U (1970) Cleavage of structural proteins during assembly of bacteriophage T4. Nature 227:680–685

    PubMed  Google Scholar 

  • Laing K, Holland J, Bonilla S, Cunningham C, Secombes C (2001) Cloning and sequencing of caspase 6 in rainbow trout (Oncorhynchus mykiss), and its expression under conditions known to induce apoptosis. Dev Comp Immunol 25:303–312

    Article  CAS  PubMed  Google Scholar 

  • Locksley R, Kileen N, Lenardo M (2001) The TNF and TNFR superfamilies: integrating mammalian biology. Cell 104:487–501

    Article  CAS  PubMed  Google Scholar 

  • Mallet S, Barclay A (1991) A new superfamily of cell surface proteins related to the nerve growth factor receptor. Immunol Today 12:220–223

    PubMed  Google Scholar 

  • Masumoto J, Zhou W, Chen FF, Su F, Kuwada Y, Hidaka E, Katsuyama T, Sagara J, Taniguchi S, Ngo-Hazelett P, Postlethwait JH, Nunez G, Inohara N (2003) Caspy, a zebrafish caspase, activated by ASC oligomerization is required for pharyngeal arch development. J Biol Chem 278:4268–4276

    Article  CAS  PubMed  Google Scholar 

  • McDonald E III, Chui P, Martelli P, Dicker D, El-Deiry W (2001) Death domain mutagenesis of KILLER/DR5 reveals residues critical for apoptotic signaling. J Biol Chem 276:14939–14945

    Article  CAS  PubMed  Google Scholar 

  • Miller NW, Chinchar VG, Clem LW (1994a) Development of leukocyte cell lines from the channel catfish (Ictalurus punctatus). J Tissue Cult Methods 16:1–7

    Google Scholar 

  • Miller NW, Rycyzyn MA, Wilson MR, Warr GW, Naftel JP, Clem LW (1994b) Development and characterization of channel catfish long term B cell lines. J Immunol 152:2180–2189

    CAS  PubMed  Google Scholar 

  • Nagata S, Golstein P (1995) The Fas death factor. Science 267:1449–1456

    CAS  PubMed  Google Scholar 

  • Nicholson D, Thornberry N (1997) Caspases: killer proteases. Trends Biochem Sci 22:299–306

    Article  CAS  PubMed  Google Scholar 

  • Nielsen H, Engelbrecht J, Brunak S, von Heijne G (1997) Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Protein Eng 10:1–6

    Article  CAS  Google Scholar 

  • Park H, Zhou H, Bengten E, Wilson M, Chinchar VG, Clem LW, Miller NW (2002) Activation of channel catfish (Ictalurus punctatus) T cells involves NFAT-like transcription factors. Dev Comp Immunol 26:775–784

    Article  CAS  PubMed  Google Scholar 

  • Ruoslahti E (1996) RGD and other recognition sites for integrins. Dev Biol 12:696–715

    Google Scholar 

  • Rycyzyn M, Wilson M, Bengten E, Warr G, Clem LW, Miller NW (1998) Mitogen and growth factor-induced activation of a STAT-like molecule in channel catfish lymphoid cells. Mol Immunol 35(2):127–136

    Article  CAS  PubMed  Google Scholar 

  • Saeki K, You A, Kato M, Miyazonon K, Yazaki Y, Takaku F (1997) Cell density-dependent apoptosis in HL-60 cells, which is mediated by an unknown soluble factor, is inhibited by transforming growth factor β1 and overexpression of Bcl-2. J Biol Chem 272:20003–20010

    Article  CAS  PubMed  Google Scholar 

  • Sambrook J, Russel DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory, New York (Plainview)

    Google Scholar 

  • Shen L, Stuge TB, Zhou H, Khayat M, Barker KS, Quiniou S, Wilson M, Bengten E, Chinchar VG, Clem LW, Miller NW (2002) Channel catfish cytotoxic cells: a mini-review. Dev Comp Immunol 26:141–149

    Article  CAS  PubMed  Google Scholar 

  • Shen L, Stuge T, Bengten E, Wilson M, Chinchar VG, Naftel J, Bernanke J, Clem LW, Miller NW (2004) Identification and characterization of clonal NK-like cells from channel catfish (Ictalurus punctatus). Dev Comp Immunol 28:139–152

    Article  CAS  PubMed  Google Scholar 

  • Smith C, Farrah T, Goodwin R (1994) The TNF receptor superfamily of cellular and viral proteins: activation, costimulation, and death. Cell 76:959–962

    Article  CAS  PubMed  Google Scholar 

  • Sprang S (1990) The divergent receptors for TNF. Trends Biochem Sci 15:366–368

    Article  CAS  PubMed  Google Scholar 

  • Stuge TB, Wilson MR, Zhou H, Barker KS, Bengten E, Chinchar G, Miller NW, Clem LW (2000) Development and analysis of various clonal alloantigen-dependent cytotoxic cell lines from channel catfish. J Immunol 164:2971–2977

    CAS  PubMed  Google Scholar 

  • Taniguichi T (1988) Regulation of cytokine gene expression. Annu Rev Immunol 6:439–464

    Article  PubMed  Google Scholar 

  • Tibbetts MD, Zheng L, Lenardo MJ (2003) The death effector domain protein family: regulators of cellular homeostasis. Nat Immunol 4:404–409

    Article  CAS  PubMed  Google Scholar 

  • Wallach D, Varfolomeev E, Malinin N, Goltsev Y, Kovalenko A, Boldin M (1999) Tumor necrosis factor receptor and Fas signaling mechanisms. Annu Rev Immunol 17:331–367

    Article  CAS  PubMed  Google Scholar 

  • Weber CH, Vincenz C (2001) The death domain superfamily: a tale of two interfaces? Trends Biochem Sci 26:475–481

    Article  CAS  PubMed  Google Scholar 

  • Wiley SR, Schooley K, Smolak PJ, Din WS, Huang CP, Nicholl JK, Sutherland GR, Smith TD, Rauch C, Smith CA (1995) Identification and characterization of a new member of the TNF family that induces apoptosis. Immunity 3:673–682

    Article  CAS  PubMed  Google Scholar 

  • Yoshida S, Stuge T, Miller NW, Clem LW (1995) Phylogeny of lymphocyte heterogeneity: cytotoxic activity of channel catfish peripheral blood leukocytes directed against allogeneic targets. Dev Comp Immunol 19:71–77

    Article  CAS  PubMed  Google Scholar 

  • Zhou H, Stuge T, Miller NW, Bengten E, Naftel J, Bernarke J, Chinchar VG, Clem LW, Wilson M (2001) Heterogeneity of channel catfish CTL with respect to target recognition and cytotoxic mechanisms employed. J Immunol 167:1325–1332

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We wish to thank Noel Hawke (Lineberger Cancer Center, University of North Carolina Medical School) for contributing the catfish FADD sequence. This work was supported by grants from the US Department of Agriculture (NRI/CGP 99-35204-7944 and 2002-35204-12211) and the National Institutes of Health (ROI-AI-19530). Nucleotide sequencing was performed in collaboration with Dr. Greg Warr and Darlene Middleton (Medical University of South Carolina).

Author information

Authors and Affiliations

  1. Department of Microbiology, University of Mississippi Medical Center, 2500 North State Street, Jackson, MS, 39216, USA

    Scott Long, Melanie Wilson, Eva Bengtén, L. William Clem, Norman W. Miller & V. Gregory Chinchar

Authors
  1. Scott Long
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Melanie Wilson
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Eva Bengtén
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. L. William Clem
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Norman W. Miller
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. V. Gregory Chinchar
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to V. Gregory Chinchar.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Long, S., Wilson, M., Bengtén, E. et al. Identification and characterization of a FasL-like protein and cDNAs encoding the channel catfish death-inducing signaling complex. Immunogenetics 56, 518–530 (2004). https://doi.org/10.1007/s00251-004-0701-2

Download citation

  • Received:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00251-004-0701-2

Keywords

Search

Navigation