Expression of an inducible nitric oxide synthase gene in rainbow trout Oncorhynchus mykiss

K J Laing, L J Hardie, W Aartsen, P S Grabowski, C J Secombes

Research output: Contribution to journalArticle

79 Citations (Scopus)

Abstract

Using oligonucleotide primers based on mammalian nitric oxide synthases (SOS), expression of an inducible NOS (iNOS) gene was detected in head kidney and gill tissue of bacterially-challenged rainbow trout. Three overlapping fragments were amplified by RT-PCR and used to construct a contiguous sequence of 1410 bp, with high nucleotide homology to iNOS in birds (61%) and mammals (57-59%). The nucleotide sequence translated in one reading frame to produce a partial peptide containing 470 amino acids, with 69-71% amino acid homology with mammalian iNOS, 81% homology with chicken iNOS and 85% homology with a partial (492 bp) goldfish iNOS sequence. In vitro stimulation of head kidney macrophages with LPS also induced expression of the trout iNOS RNA. with optimal expression seen using 20 50 mu g/ml LPS at 2 h to 6 h post-stimulation. The evolutionary and functional significance of the trout iNOS sequence are discussed. (C) 1999 Elsevier Science Ltd. All rights reserved.

Original languageEnglish
Pages (from-to)71-85
Number of pages15
JournalDevelopmental and Comparative Immunology
Volume23
Publication statusPublished - 1999

Keywords

  • fish
  • macrophages
  • iNOS
  • sequence
  • RT-PCR
  • expression
  • MACROPHAGE CELL-LINE
  • L-ARGININE
  • MOLECULAR-CLONING
  • ACTIVATED MACROPHAGES
  • ICTALURUS-PUNCTATUS
  • ENDOTHELIAL-CELLS
  • CHANNEL CATFISH
  • MESSENGER-RNA
  • CDNA CLONING
  • CALMODULIN

Cite this

Expression of an inducible nitric oxide synthase gene in rainbow trout Oncorhynchus mykiss. / Laing, K J ; Hardie, L J ; Aartsen, W ; Grabowski, P S ; Secombes, C J .

In: Developmental and Comparative Immunology, Vol. 23, 1999, p. 71-85.

Research output: Contribution to journalArticle

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abstract = "Using oligonucleotide primers based on mammalian nitric oxide synthases (SOS), expression of an inducible NOS (iNOS) gene was detected in head kidney and gill tissue of bacterially-challenged rainbow trout. Three overlapping fragments were amplified by RT-PCR and used to construct a contiguous sequence of 1410 bp, with high nucleotide homology to iNOS in birds (61{\%}) and mammals (57-59{\%}). The nucleotide sequence translated in one reading frame to produce a partial peptide containing 470 amino acids, with 69-71{\%} amino acid homology with mammalian iNOS, 81{\%} homology with chicken iNOS and 85{\%} homology with a partial (492 bp) goldfish iNOS sequence. In vitro stimulation of head kidney macrophages with LPS also induced expression of the trout iNOS RNA. with optimal expression seen using 20 50 mu g/ml LPS at 2 h to 6 h post-stimulation. The evolutionary and functional significance of the trout iNOS sequence are discussed. (C) 1999 Elsevier Science Ltd. All rights reserved.",
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AU - Secombes, C J

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N2 - Using oligonucleotide primers based on mammalian nitric oxide synthases (SOS), expression of an inducible NOS (iNOS) gene was detected in head kidney and gill tissue of bacterially-challenged rainbow trout. Three overlapping fragments were amplified by RT-PCR and used to construct a contiguous sequence of 1410 bp, with high nucleotide homology to iNOS in birds (61%) and mammals (57-59%). The nucleotide sequence translated in one reading frame to produce a partial peptide containing 470 amino acids, with 69-71% amino acid homology with mammalian iNOS, 81% homology with chicken iNOS and 85% homology with a partial (492 bp) goldfish iNOS sequence. In vitro stimulation of head kidney macrophages with LPS also induced expression of the trout iNOS RNA. with optimal expression seen using 20 50 mu g/ml LPS at 2 h to 6 h post-stimulation. The evolutionary and functional significance of the trout iNOS sequence are discussed. (C) 1999 Elsevier Science Ltd. All rights reserved.

AB - Using oligonucleotide primers based on mammalian nitric oxide synthases (SOS), expression of an inducible NOS (iNOS) gene was detected in head kidney and gill tissue of bacterially-challenged rainbow trout. Three overlapping fragments were amplified by RT-PCR and used to construct a contiguous sequence of 1410 bp, with high nucleotide homology to iNOS in birds (61%) and mammals (57-59%). The nucleotide sequence translated in one reading frame to produce a partial peptide containing 470 amino acids, with 69-71% amino acid homology with mammalian iNOS, 81% homology with chicken iNOS and 85% homology with a partial (492 bp) goldfish iNOS sequence. In vitro stimulation of head kidney macrophages with LPS also induced expression of the trout iNOS RNA. with optimal expression seen using 20 50 mu g/ml LPS at 2 h to 6 h post-stimulation. The evolutionary and functional significance of the trout iNOS sequence are discussed. (C) 1999 Elsevier Science Ltd. All rights reserved.

KW - fish

KW - macrophages

KW - iNOS

KW - sequence

KW - RT-PCR

KW - expression

KW - MACROPHAGE CELL-LINE

KW - L-ARGININE

KW - MOLECULAR-CLONING

KW - ACTIVATED MACROPHAGES

KW - ICTALURUS-PUNCTATUS

KW - ENDOTHELIAL-CELLS

KW - CHANNEL CATFISH

KW - MESSENGER-RNA

KW - CDNA CLONING

KW - CALMODULIN

M3 - Article

VL - 23

SP - 71

EP - 85

JO - Developmental and Comparative Immunology

JF - Developmental and Comparative Immunology

SN - 0145-305X

ER -