Analysis and characterisation of IL-1 beta processing in rainbow trout, Oncorhynchus mykiss

S H Hong, Jun Zou, Bertrand Collet, N C Bols, Christopher John Secombes

Research output: Contribution to journalArticle

34 Citations (Scopus)

Abstract

Mammalian IL-1beta is produced as a biologically inactive 31 kDa precursor, which is converted to the active 18 kDa form by proteolytic processing. Synthesis and processing of native piscine IL-1beta is poorly understood. In the present study, the native IL-1beta precursor or mature peptides were detected at sizes of approx. 29 kDa and 24 kDa in cell lysates of a rainbow trout macrophage cell line RTS-11, with or without LPS stimulation, by Western blot analysis using a polyclonal antibody against the putative trout mature IL-1beta (rmIL-1beta) produced in Escherichia coli. Processing of the 29 kDa precursor into a 24 kDa mature peptide was confirmed by analysis of such proteins using a monoclonal conjugate (Ni-NTA-HRP) against 6 histidines in lysates of the RTS-11 cells transfected with an expression plasmid containing the IL-1beta precursor molecule tagged with 6 histidines at its C terminus. Only the recombinant mature 24 kDa) IL-1beta/HIS protein was purified from the culture supernatants of the transfected cells, indicating the molecule is cleaved to be secreted. These findings strongly suggest that the trout IL-1beta molecule is processed in trout macrophages in an analogous way to the situation with mammalian IL-1beta despite the lack of clear ICE cut site. (C) 2003 Elsevier Ltd. All rights reserved.

Original languageEnglish
Pages (from-to)453-459
Number of pages7
JournalFish & Shellfish Immunology
Volume16
Issue number3
Early online date28 Sep 2003
DOIs
Publication statusPublished - Mar 2004

Keywords

  • interleukin-1 beta
  • processing
  • ICE-independent
  • rainbow trout
  • converting-enzyme
  • cysteine protease
  • cell-line
  • IL-1 beta
  • interleukin-1-beta
  • precursor
  • monocytes
  • cleavage
  • produce

Cite this

Analysis and characterisation of IL-1 beta processing in rainbow trout, Oncorhynchus mykiss. / Hong, S H ; Zou, Jun; Collet, Bertrand; Bols, N C ; Secombes, Christopher John.

In: Fish & Shellfish Immunology, Vol. 16, No. 3, 03.2004, p. 453-459.

Research output: Contribution to journalArticle

Hong, S H ; Zou, Jun ; Collet, Bertrand ; Bols, N C ; Secombes, Christopher John. / Analysis and characterisation of IL-1 beta processing in rainbow trout, Oncorhynchus mykiss. In: Fish & Shellfish Immunology. 2004 ; Vol. 16, No. 3. pp. 453-459.
@article{01faa761a410442284809083d3cf0538,
title = "Analysis and characterisation of IL-1 beta processing in rainbow trout, Oncorhynchus mykiss",
abstract = "Mammalian IL-1beta is produced as a biologically inactive 31 kDa precursor, which is converted to the active 18 kDa form by proteolytic processing. Synthesis and processing of native piscine IL-1beta is poorly understood. In the present study, the native IL-1beta precursor or mature peptides were detected at sizes of approx. 29 kDa and 24 kDa in cell lysates of a rainbow trout macrophage cell line RTS-11, with or without LPS stimulation, by Western blot analysis using a polyclonal antibody against the putative trout mature IL-1beta (rmIL-1beta) produced in Escherichia coli. Processing of the 29 kDa precursor into a 24 kDa mature peptide was confirmed by analysis of such proteins using a monoclonal conjugate (Ni-NTA-HRP) against 6 histidines in lysates of the RTS-11 cells transfected with an expression plasmid containing the IL-1beta precursor molecule tagged with 6 histidines at its C terminus. Only the recombinant mature 24 kDa) IL-1beta/HIS protein was purified from the culture supernatants of the transfected cells, indicating the molecule is cleaved to be secreted. These findings strongly suggest that the trout IL-1beta molecule is processed in trout macrophages in an analogous way to the situation with mammalian IL-1beta despite the lack of clear ICE cut site. (C) 2003 Elsevier Ltd. All rights reserved.",
keywords = "interleukin-1 beta, processing, ICE-independent, rainbow trout, converting-enzyme, cysteine protease, cell-line, IL-1 beta, interleukin-1-beta, precursor, monocytes, cleavage, produce",
author = "Hong, {S H} and Jun Zou and Bertrand Collet and Bols, {N C} and Secombes, {Christopher John}",
year = "2004",
month = "3",
doi = "10.1016/j.fsi.2003.08.002",
language = "English",
volume = "16",
pages = "453--459",
journal = "Fish & Shellfish Immunology",
issn = "1050-4648",
publisher = "ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD",
number = "3",

}

TY - JOUR

T1 - Analysis and characterisation of IL-1 beta processing in rainbow trout, Oncorhynchus mykiss

AU - Hong, S H

AU - Zou, Jun

AU - Collet, Bertrand

AU - Bols, N C

AU - Secombes, Christopher John

PY - 2004/3

Y1 - 2004/3

N2 - Mammalian IL-1beta is produced as a biologically inactive 31 kDa precursor, which is converted to the active 18 kDa form by proteolytic processing. Synthesis and processing of native piscine IL-1beta is poorly understood. In the present study, the native IL-1beta precursor or mature peptides were detected at sizes of approx. 29 kDa and 24 kDa in cell lysates of a rainbow trout macrophage cell line RTS-11, with or without LPS stimulation, by Western blot analysis using a polyclonal antibody against the putative trout mature IL-1beta (rmIL-1beta) produced in Escherichia coli. Processing of the 29 kDa precursor into a 24 kDa mature peptide was confirmed by analysis of such proteins using a monoclonal conjugate (Ni-NTA-HRP) against 6 histidines in lysates of the RTS-11 cells transfected with an expression plasmid containing the IL-1beta precursor molecule tagged with 6 histidines at its C terminus. Only the recombinant mature 24 kDa) IL-1beta/HIS protein was purified from the culture supernatants of the transfected cells, indicating the molecule is cleaved to be secreted. These findings strongly suggest that the trout IL-1beta molecule is processed in trout macrophages in an analogous way to the situation with mammalian IL-1beta despite the lack of clear ICE cut site. (C) 2003 Elsevier Ltd. All rights reserved.

AB - Mammalian IL-1beta is produced as a biologically inactive 31 kDa precursor, which is converted to the active 18 kDa form by proteolytic processing. Synthesis and processing of native piscine IL-1beta is poorly understood. In the present study, the native IL-1beta precursor or mature peptides were detected at sizes of approx. 29 kDa and 24 kDa in cell lysates of a rainbow trout macrophage cell line RTS-11, with or without LPS stimulation, by Western blot analysis using a polyclonal antibody against the putative trout mature IL-1beta (rmIL-1beta) produced in Escherichia coli. Processing of the 29 kDa precursor into a 24 kDa mature peptide was confirmed by analysis of such proteins using a monoclonal conjugate (Ni-NTA-HRP) against 6 histidines in lysates of the RTS-11 cells transfected with an expression plasmid containing the IL-1beta precursor molecule tagged with 6 histidines at its C terminus. Only the recombinant mature 24 kDa) IL-1beta/HIS protein was purified from the culture supernatants of the transfected cells, indicating the molecule is cleaved to be secreted. These findings strongly suggest that the trout IL-1beta molecule is processed in trout macrophages in an analogous way to the situation with mammalian IL-1beta despite the lack of clear ICE cut site. (C) 2003 Elsevier Ltd. All rights reserved.

KW - interleukin-1 beta

KW - processing

KW - ICE-independent

KW - rainbow trout

KW - converting-enzyme

KW - cysteine protease

KW - cell-line

KW - IL-1 beta

KW - interleukin-1-beta

KW - precursor

KW - monocytes

KW - cleavage

KW - produce

U2 - 10.1016/j.fsi.2003.08.002

DO - 10.1016/j.fsi.2003.08.002

M3 - Article

VL - 16

SP - 453

EP - 459

JO - Fish & Shellfish Immunology

JF - Fish & Shellfish Immunology

SN - 1050-4648

IS - 3

ER -