Effect of growth rate on transcriptomic responses to immune stimulation in wild-type, domesticated, and GH-transgenic coho salmon

Abstract

Background: Transcriptomic responses to immune stimulation were investigated in coho salmon (Oncorhynchus kisutch) with distinct growth phenotypes. Wild-type fish were contrasted to strains with accelerated growth arising either from selective breeding (i.e. domestication) or genetic modification. Such distinct routes to accelerated growth may have unique implications for relationships and/or trade-offs between growth and immune function.

Results: RNA-Seq was performed on liver and head kidney in four 'growth response groups' injected with polyinosinic-polycytidylic acid (Poly I:C; viral mimic), peptidoglycan (PGN; bacterial mimic) or PBS (control). These groups were: 1) 'W': wild-type, 2) 'TF': growth hormone (GH) transgenic salmon with ~ 3-fold higher growth-rate than W, 3) 'TR': GH transgenic fish ration restricted to possess a growth-rate equal to W, and 4) 'D': domesticated non-transgenic fish showing growth-rate intermediate to W and TF. D and TF showed a higher similarity in transcriptomic response compared to W and TR. Several immune genes showed constitutive expression differences among growth response groups, including perforin 1 and C-C motif chemokine 19-like. Among the affected immune pathways, most were up-regulated by Poly I:C and PGN. In response to PGN, the c-type lectin receptor signalling pathway responded uniquely in TF and TR. In response to stimulation with both immune mimics, TR responded more strongly than other groups. Further, group-specific pathway responses to PGN stimulation included NOD-like receptor signalling in W and platelet activation in TR. TF consistently showed the most attenuated immune response relative to W, and more DEGs were apparent in TR than TF and D relative to W, suggesting that a non-satiating ration coupled with elevated circulating GH levels may cause TR to possess enhanced immune capabilities. Alternatively, TF and D salmon are prevented from acquiring the same level of immune response as TR due to direction of energy to high overall somatic growth. Further study of the effects of ration restriction in growth-modified fishes is warranted.

Conclusions: These findings improve our understanding of the pleiotropic effects of growth modification on the immunological responses of fish, revealing unique immune pathway responses depending on the mechanism of growth acceleration and nutritional availability.

Keywords: Coho salmon; Growth; Growth hormone; Immunity; PGN; Pleiotropy; Poly I:C; Selective breeding; Transcriptomics; Transgenesis.

Fig. 1

Principal component analysis (PCA) of all reads from RNA-Seq analysis for a) Head kidney and b) Liver treated with PBS, PGN and Poly I:C for. PBS, phosphate-buffered saline; PGN, peptidoglycan; Poly I:C, polyinosinic-polycytidylic acid. W, non-transgenic (wild-type) coho salmon on a full satiation ration; TF, GH transgenic coho salmon on a full satiation ration; TR, GH transgenic coho salmon on restricted ration equal to that consumed by W; D, domesticated coho salmon on a full satiation ration

Fig. 2

Bioinformatic analysis plan for the study. a) Pairwise assessment within each group, comparing immune-stimulated fish to their respective group treated with PBS, peptidoglycan (PGN) and Poly I:C. b) number of differentially expressed genes (DEGs) identified in the study by two different statistical normalized methods (Baggerley’s and DESeq2) treated with PBS, c0 peptidoglycan (PGN), and d) Poly I:C. Numbers refer to DEGs displaying a fold-change ≥3 among fish groups, with a normalized false discovery rate (FDR) P-value correction < 0.01). See Fig. 1 legend for abbreviations

Fig. 3

Heatmap of differentially expressed genes (DEGs) from comparisons among PBS-treated groups a) head kidney and b) liver. A star within cells refers to DEGs determined by the criteria of fold-change ≥3, and a normalized false discovery rate (FDR) P-value correction < 0.01. See Fig. 1 legend for abbreviations

Fig. 4

a) Number of differentially expressed gene (DEG) shared among comparisons within the fish groups (TF, TR, W, D) treated with immune stimulants Poly I:C, relative to each respective fish group treated with PBS, for both head kidney and liver. b) Heatmap for all significant differentially expressed gene (DEG) for comparison within fish groups treated with immune stimulants compared the same group treated with PBS for both head kidney and liver. Hierarchical clustering analysis was performed by MeV (ver. 4.9; https://sourceforge.net/projects/mev-tm4/files/mev-tm4/). A star within cells refers to DEGs determined by the criteria of fold-change ≥3, and a normalized false discovery rate (FDR) P-value correction < 0.01. See Fig. 1 legend for abbreviations

Fig. 5

Heatmap for immune-related KEGG pathway-annotated differentially expressed gene (DEG) for both a) head kidney and b) liver for each comparison within the fish groups treated with PGN and Poly I:C. Hierarchical clustering analysis was performed by MeV (ver. 4.9; https://sourceforge.net/projects/mev-tm4/files/mev-tm4/). A star within cells refers to DEGs determined by the criteria of fold-change ≥3, and a normalized false discovery rate (FDR) P-value correction < 0.01. See Fig. 1 legend for abbreviations

Fig. 6

Diagram illustrating relationships of significant immune-related KEGG pathway differentially expressed gene (DEG) identified within the fish groups treated with a) bacterial and b) viral mimics. The number and size of circles within the figure corresponds to the number of DEG in the affected pathway. Overlapping circles represent shared responses. The color of each circle refers to the value of gene expression according to the fold change. See Fig. 1 legend for abbreviations