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Interpretation of the Genotype by Tissue Interactions of Four Genes (AFP1, CIRP, YB-1, and HMGB1) in Takifugu rubripes Under Different Low-Temperature Conditions

ABSTRACT: Background: The differential expression of the same gene in different tissues could be due to the genotype effect, tissue effect, and/or genotype × tissue interactions. However, the genetic mechanisms responsible for this differential expression have not been reported to date. Methods: Four resistance genes to low temperature, the genes for antifreeze protein (AFP), cold induced RNA binding protein (CIRP), high mobility group protein box-1 (HMGB1), and Y-box binding protein (YB-1), were measured by PCR in the liver, spleen, kidney, brain, heart, intestine, muscle, gonad, and skin of Takifugu rubripes cultured under different temperature conditions (18, 13, 8, and 5°C). Split-split-plot analysis of variance, additive main effects and multiplicative interaction (AMMI) and genotype main effects and genotype × environment interaction (GGE) biplot analysis were used to evaluate the effects of genotype × tissue interactions on gene expression. Results: The results of split-split-plot analysis of variance showed that water temperature has a significant effect on the expression of T. rubripes cold resistant genes, while tissue × gene interaction has a highly significant effect on it. AMMI analysis showed that the contributions of genotype, tissue, and genotype × tissue interactions to the total variation in gene expression followed two trends: 1) as temperature decreased, the gene effect increased gradually and the genotype × tissue interaction decreased gradually; 2) the gene effect at 18 and 13°C was significantly lower than that at 8 and 5°C, while the interaction at 18 and 13°C was significantly higher than that at 8 and 5°C. GGE analysis showed that: at all temperatures except 8°C, the expression rankings of all four genes were highly positively correlated in all tissues except muscle; the expression stability of the genes was the same at 18°C/13°C and at 8°C/5°C; and AFP1 showed the best expression and stability among the four genes. Conclusion: 8°C/5°C as the suitable temperature for such experiments for T. rubripes. Among the four antifreeze genes evaluated in this study, AFP1 had the best expression and stability.

SUBMITTER: Wang X

PROVIDER: S-EPMC9280165 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:DNA methylation has frequently been implicated in sex determination and differentiation in teleost species. In order to detect the DNA methylation patterns established during sexual differentiation in tiger pufferfish T. rubripes, we performed comprehensive whole genome methylation sequencing and analyses of the gonads of male, female, and pseudo male. We obtained a total of 33.12, 32.44, and 31.60 Gb clean data for male, female, and pseudo male, with a sequencing depth of 66.44×, 60.47× and 54.86×, respectively. The methylation level of cytosine (C) residues in the genomic DNA from gonads was 11.016%, 10.428%, and 11.083% in male, female, and pseudo male, respectively. More than 65% of C methylation was at CpG sites, and less than 1% was at CHG and CHH sites. In each regulatory element, there were low methylation levels on both sides of the transcription start site, and higher methylation levels in exons, introns, and downstream of genes. The highest mCpG was on chromosome 8 and the lowest mCpG was on chromosome 5. Comparisons of whole-genome DNA methylation between pairs of samples revealed that there were 3,173 differentially methylated regions (DMRs) between female and male, and 3,037 DMRs between male and pseudo male, corresponding to 0.232% and 0.223% of the length of the genome, respectively. There were only 1,635 DMRs between female and pseudo male, representing 0.127% of the length of the genome. A number of differentially methylated genes (DMGs) related to sex determination and differentiation were selected, such as amhr2 and pfcyp19a. After Bisulfite Sequencing PCR (BSP) verification, amhr2 was exhibited low methylation level in normal males and pseudo male, and high methylation level in normal females but pfcyp19a showed low methylation level in normal females and high methylation level in normal males and pseudo males. These results provide information about the molecular epigenetic mechanisms of DNA methylation during low-temperature induced masculinization of tiger pufferfish, and increase our understanding of the mechanisms of sex determination and differentiation in this important aquaculture fish species.

Project description:BackgroundGuanine protein-coupled receptors (GPCRs) constitute a eukaryotic transmembrane protein family and function as "molecular switches" in the second messenger cascades and are found in all organisms between yeast and humans. They form the single, biggest drug-target family due to their versatility of action and their role in several physiological functions, being active players in detecting the presence of light, a variety of smells and tastes, amino acids, nucleotides, lipids, chemicals etc. in the environment of the cell. Comparative genomic studies on model organisms provide information on target receptors in humans and their function. The Japanese teleost Fugu has been identified as one of the smallest vertebrate genomes and a compact model to study the human genome, owing to the great similarity in its gene repertoire with that of human and other vertebrates. Thus the characterization of the GPCRs of Fugu would provide insights to the evolution of the vertebrate genome.ResultsWe classified the GPCRs in the Fugu genome and our analysis of its 316 membrane-bound receptors, available on the public databases as well as from literature, detected 298 GPCRs that were grouped into five main families according to the GRAFS classification system (namely, Glutamate, Rhodopsin, Adhesion, Frizzled and Secretin). We also identified 18 other GPCRs that could not be grouped under the GRAFS family and hence were classified as 'Other 7TM' receptors. On comparison of the GPCR information from the Fugu genome with those in the human and chicken genomes, we detected 96.83% (306/316) and 96.51% (305/316) orthology in GPCRs among the Fugu-human genomes and Fugu-chicken genomes, respectively.ConclusionsThis study reveals the position of pisces in vertebrate evolution from the GPCR perspective. Fugu can act as a reference model for the human genome for other protein families as well, going by the high orthology observed for GPCRs between Fugu and human. The evolutionary comparison of GPCR sequences between key vertebrate classes of mammals, birds and fish will help in identifying key functional residues and motifs so as to fill in the blanks in the evolution of GPCRs in vertebrates.

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Interpretation of the Genotype by Tissue Interactions of Four Genes (AFP1, CIRP, YB-1, and HMGB1) in Takifugu rubripes Under Different Low-Temperature Conditions

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