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Molecular cloning of double-stranded RNA inducible Mx genes from Atlantic salmon (Salmo salar L.)
B Robertsen1, G Trobridge, J A Leong
1Norwegian College of Fishery Science, University of Tromsø, Norway.
Abstract:
Mx proteins are induced by type I interferons in mice and humans and inhibit the replication of several negative-stranded RNA viruses. In this work Mx genes in Atlantic salmon were studied using the double stranded RNA, polyinosinic: polycytidylic acid (poly I:C), to induce interferon production. Northern blot analysis showed Mx mRNA in liver, head kidney and gills 2 days after poly I:C injection of fish, but not in untreated fish or fish injected with saline or LPS. Mx transcripts of 2.4 and 1.9 kb were detected in the liver. By screening of a cDNA library, three different full length Mx cDNA clones, ASMx1, ASMx2 and ASMx3, were identified and sequenced. The deduced ASMx proteins all contain 623 amino acids and show the tripartite GTP-binding motif typical of vertebrate Mx proteins. ASMx1 and ASMx2 may represent alleles of the same gene whereas ASMx3 represents a different gene. The deduced ASMx proteins showed 96 to 98% sequence identity with rainbow trout Mx1 and Mx3 and about 88% identity with rainbow trout Mx2 protein.
Insights
Atlantic salmon Mx genes were investigated following interferon induction. Three Mx cDNA clones were identified, revealing insights into fish antiviral defenses and their relation to rainbow trout Mx proteins.
Area of Science:
- Immunology
- Molecular Biology
- Aquaculture
Background:
- Mx proteins are crucial antiviral effectors induced by type I interferons in vertebrates.
- Understanding Mx gene function in fish is vital for aquaculture health and disease resistance.
Purpose of the Study:
- To characterize Mx genes in Atlantic salmon (Salmo salar).
- To investigate the induction of Mx gene expression by polyinosinic:polycytidylic acid (poly I:C).
- To identify and sequence Atlantic salmon Mx cDNA clones.
Main Methods:
- Atlantic salmon were injected with poly I:C to induce interferon production.
- Mx mRNA expression was analyzed using Northern blot in various tissues.
- Full-length Mx cDNA clones were isolated by screening a cDNA library and sequenced.
Main Results:
- Mx mRNA was detected in liver, head kidney, and gills post-poly I:C injection.
- Two transcript sizes (2.4 and 1.9 kb) were observed in salmon liver.
- Three Atlantic salmon Mx (ASMx) cDNA clones (ASMx1, ASMx2, ASMx3) were identified, encoding proteins with a conserved GTP-binding motif.
- ASMx1 and ASMx2 likely represent alleles, while ASMx3 represents a distinct gene.
- ASMx proteins exhibited high sequence identity to rainbow trout Mx proteins.
Conclusions:
- Poly I:C effectively induces Mx gene expression in Atlantic salmon.
- Atlantic salmon possess multiple Mx genes, including potential alleles.
- The identified Mx proteins share conserved structural features and sequence homology with related fish species, suggesting a conserved antiviral role.