HSP70 gene expression responses to the temperature stress in pufferfish (Takifugu rubripes).

In this study, we isolated and characterized HSP70 cDNA from pufferfish (Takifugu rubripes). The 3053 bp full-length TrHSP70 sequence consisted of a 167 bp 5'-UTR (untranslated region), a 2535 bp open reading frame, and a 351 bp 3'-UTR. BLAST analysis revealed that the TrHSP70 shared high similarity with HSP70 sequences in other species. In our study, we set 3 experimental groups as H1 group (20 °C), H2 group (24 °C), and H3 group (28 °C) for checking the expression level of TrHSP70 in T. rubripes. Tissue-specific gene expression results showed that TrHSP70 had higher expression in the intestines than other tissues of the T. rubripes by RT-qPCR. In the experimental group, we found that the expression of TrHSP70 was upregulated in different tissues in the H3 group. The results show that TrHSP70 is a constitutively expressed gene, which plays an important role in maintaining normal physiological function and coping with stress.

Quantitative proteomic analysis in serum of Takifugu rubripes infected with Cryptocaryon irritans.

Cryptocaryon irritans can cause cryptocaryonosis (white spot disease) in marine fish but the pathogenesis of the disease is unclear. In this work, we used high-throughput proteomics to identify differentially expressed proteins in the serum of Takifugu rubripes challenged with C. irritans. By using quantitative proteomic assays combined with Tandem Mass Tag-labeled quantitative proteomic analysis, we identified a total of 2088 differentially abundant proteins (1706 proteins were quantified, p < 0.05, fold-change threshold ≥ 2), including 21 up-regulated and 44 down-regulated. Combined with STRING-based functional analysis, we ultimately obtained eight proteins including glucokinase-like, integrin beta-1-like isoform X2, H4, H2A.V, histone H1-like, histone H2AX-like, histone H2B 1/2-like and myosin-9 isoform X1, which could be considered as potential biomarkers for T. rubripes immune responses. Eight proteins that were selected to validate significant differentially expressed genes at the proteomic level were consistent with qPCR at the transcriptomic level. The proteins identified in our work may serve as candidates for elucidating the molecular mechanism of cryptocaryonosis in T. rubripes. Our collective findings could provide new insights into searching for disease-specific targets and biomarkers, which may be effective indicators of C. irritans infection in T. rubripes.

Characterization and expression analysis of the C8α and C9 terminal complement components from pufferfish (Takifugu rubripes).

C8α and C9 mediate the membrane attack complex formation and bacterial lysis and are important components in the complement system. The cDNA sequences of the C8α and C9 genes were cloned from Takifugu rubripes. The full-length cDNA of Tr-C8α was 1893 bp and included a 5'-UTR of 69 bp and 3'-UTR of 83 bp. The full-length cDNA of Tr-C9 was 2083 bp and included a 5'-UTR of 72 bp and 3'-UTR of 250 bp. The expression of Tr-C8α and Tr-C9 was detected in newly fertilized eggs of T. rubripes. The expression of these two genes was at a higher level in the liver than in other tissues tested. After lipopolysaccharide (LPS) challenge, the gene expression of Tr-C8α and Tr-C9 increased more significantly in the liver. With these combined results, we further understood how Tr-C8α and Tr-C9 function in the innate immunity of pufferfish. Our findings could deepen the understanding of immune regulation in pufferfish.