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An ancient immunity gene duplication in Daphnia magna: RNA expression and sequence analysis of two nitric oxide synthase genes.

Labbé P, McTaggart SJ, Little TJ - Dev. Comp. Immunol. (2009)

Bottom Line: Both genes bear features commonly found in invertebrate NOS, however, the two genes differ in their rate of evolution, intraspecific polymorphism and expression level.We tested whether the more rapid evolution of NOS2 could be due to positive selection, but found the rate of amino-acid substitutions between Daphnia species to be compatible with a neutral model.A second experiment indicated that NOS transcription does not increase following exposure to Pasteuria.

View Article: PubMed Central - PubMed

Affiliation: University of Edinburgh, Institute of Evolutionary Biology, School of Biological Sciences, Ashworth Laboratory, Kings Buildings, Edinburgh, UK. Pierrick.Labbe@ed.ac.uk

ABSTRACT
NO (nitric oxide) is a highly reactive free radical gas thought to play a major role in the invertebrate immune response by harming pathogens and limiting their growth. Here we report on studies of nitric oxide synthase (NOS) genes in the crustacean Daphnia, one of the few non-insect arthropod models used to study host-pathogen interactions. While the NOS gene is found as a single copy in other invertebrates, we found two copies (NOS1 and NOS2), which a phylogenetic reconstruction showed to be the result of an ancient duplication event. Both genes bear features commonly found in invertebrate NOS, however, the two genes differ in their rate of evolution, intraspecific polymorphism and expression level. We tested whether the more rapid evolution of NOS2 could be due to positive selection, but found the rate of amino-acid substitutions between Daphnia species to be compatible with a neutral model. To associate NOS or NO activity with infection, we performed infection experiments with Daphnia magna and one of its natural pathogens (the bacterium Pasteuria ramosa). In one set of experimental infections, we supplemented D. magna with L-arginine, the NOS substrate, or with L-NAME, a NOS antagonist, and found this to result in lower and higher infection levels, respectively, which is at least compatible with the notion that NO may aid defence against Pasteuria. A second experiment indicated that NOS transcription does not increase following exposure to Pasteuria. Thus, the function of NOS in Daphnia immunity remains uncertain, but the pattern of gene duplication and subsequent divergence suggests evolution via neo- or subfunctionalization.

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Multiple alignment for conserved motives of several arthropod NOS amino-acid sequences. See text for the reference sequences of the various organisms; for reasons of print space not all the sequences used are presented but only a representative selection of the diversity (in particular as D. pulex and D. magna are relatively close, the D. pulex sequences are not shown). Conserved amino-acid in more than 90% of the sequences are black shaded, those conserved in more than 50% of the sequences are grey shaded. Binding sites to heme, tetragydrobiopterin (BH4), calmodulin, FMN, FAD pyrophosphate, FAD isoalloxazine, flavin, NADPH ribose, NADPH adenine and NADPH are underlined.
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fig3: Multiple alignment for conserved motives of several arthropod NOS amino-acid sequences. See text for the reference sequences of the various organisms; for reasons of print space not all the sequences used are presented but only a representative selection of the diversity (in particular as D. pulex and D. magna are relatively close, the D. pulex sequences are not shown). Conserved amino-acid in more than 90% of the sequences are black shaded, those conserved in more than 50% of the sequences are grey shaded. Binding sites to heme, tetragydrobiopterin (BH4), calmodulin, FMN, FAD pyrophosphate, FAD isoalloxazine, flavin, NADPH ribose, NADPH adenine and NADPH are underlined.


An ancient immunity gene duplication in Daphnia magna: RNA expression and sequence analysis of two nitric oxide synthase genes.

Labbé P, McTaggart SJ, Little TJ - Dev. Comp. Immunol. (2009)

Multiple alignment for conserved motives of several arthropod NOS amino-acid sequences. See text for the reference sequences of the various organisms; for reasons of print space not all the sequences used are presented but only a representative selection of the diversity (in particular as D. pulex and D. magna are relatively close, the D. pulex sequences are not shown). Conserved amino-acid in more than 90% of the sequences are black shaded, those conserved in more than 50% of the sequences are grey shaded. Binding sites to heme, tetragydrobiopterin (BH4), calmodulin, FMN, FAD pyrophosphate, FAD isoalloxazine, flavin, NADPH ribose, NADPH adenine and NADPH are underlined.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2724039&req=5

fig3: Multiple alignment for conserved motives of several arthropod NOS amino-acid sequences. See text for the reference sequences of the various organisms; for reasons of print space not all the sequences used are presented but only a representative selection of the diversity (in particular as D. pulex and D. magna are relatively close, the D. pulex sequences are not shown). Conserved amino-acid in more than 90% of the sequences are black shaded, those conserved in more than 50% of the sequences are grey shaded. Binding sites to heme, tetragydrobiopterin (BH4), calmodulin, FMN, FAD pyrophosphate, FAD isoalloxazine, flavin, NADPH ribose, NADPH adenine and NADPH are underlined.
Bottom Line: Both genes bear features commonly found in invertebrate NOS, however, the two genes differ in their rate of evolution, intraspecific polymorphism and expression level.We tested whether the more rapid evolution of NOS2 could be due to positive selection, but found the rate of amino-acid substitutions between Daphnia species to be compatible with a neutral model.A second experiment indicated that NOS transcription does not increase following exposure to Pasteuria.

View Article: PubMed Central - PubMed

Affiliation: University of Edinburgh, Institute of Evolutionary Biology, School of Biological Sciences, Ashworth Laboratory, Kings Buildings, Edinburgh, UK. Pierrick.Labbe@ed.ac.uk

ABSTRACT
NO (nitric oxide) is a highly reactive free radical gas thought to play a major role in the invertebrate immune response by harming pathogens and limiting their growth. Here we report on studies of nitric oxide synthase (NOS) genes in the crustacean Daphnia, one of the few non-insect arthropod models used to study host-pathogen interactions. While the NOS gene is found as a single copy in other invertebrates, we found two copies (NOS1 and NOS2), which a phylogenetic reconstruction showed to be the result of an ancient duplication event. Both genes bear features commonly found in invertebrate NOS, however, the two genes differ in their rate of evolution, intraspecific polymorphism and expression level. We tested whether the more rapid evolution of NOS2 could be due to positive selection, but found the rate of amino-acid substitutions between Daphnia species to be compatible with a neutral model. To associate NOS or NO activity with infection, we performed infection experiments with Daphnia magna and one of its natural pathogens (the bacterium Pasteuria ramosa). In one set of experimental infections, we supplemented D. magna with L-arginine, the NOS substrate, or with L-NAME, a NOS antagonist, and found this to result in lower and higher infection levels, respectively, which is at least compatible with the notion that NO may aid defence against Pasteuria. A second experiment indicated that NOS transcription does not increase following exposure to Pasteuria. Thus, the function of NOS in Daphnia immunity remains uncertain, but the pattern of gene duplication and subsequent divergence suggests evolution via neo- or subfunctionalization.

Show MeSH
Related in: MedlinePlus