Physiological Effects and Transcriptomic Analysis of sbGnRH on the Liver in Pompano (Trachinotus ovatus)

Abstract

Pompano (Trachinotus ovatus) is one of the important economic marine fishes in the south coast of China. At present, the research on the basic biology of pompano is relatively weak, which has seriously affected the development of this economic important fish. The liver is an important digestive and metabolic organ of fish which plays an important regulatory role in its growth and development. It is necessary to clarify the effects of sea bream gonadotropin releasing hormone (sbGnRH) on liver physiology and metabolic enzyme activity. The effects of sbGnRH peptides (10 ng/gbw) on the physiological and biochemical indices and metabolic enzyme activities of pompano liver were studied. It was found that after injection of 10 ng/gbw sbGnRH peptides, the contents of albumin, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, glucose, creatine kinase, iron, magnesium, aspartate aminotransferase, alanine aminotransferase and creatinine increased, while of cholesterol and calcium contents decreased. The activities of amylase, lipase, pyruvate kinase, acyl CoA oxidase, superoxide dismutase, phospholipid hydroperoxide glutathione peroxidase, catalase, glucose-6-phosphate dehydrogenase, fatty acid synthase and lipoprotein lipase increased, while the activities of malic enzyme, carnitine acyl, carnitine translocation, acetyl CoA carboxylase and malondialdehyde decreased. Three hours after the injection of different concentrations of sbGnRH peptides (0 and 10 ng/gbw), the transcriptome sequences of the two groups of livers were sequenced. After quality control and removal of some low-quality data, clean reads of 21,283,647、19,427,359、21,873,990、21,732,174、23,660,062 and 21,592,338 were obtained respectively. In this study, 99 genes were screened and identified as differentially expressed genes, including 77 up-regulated genes and 22 down-regulated genes. According to the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) pathway analyses, these pathways and the typical genes involved can be divided into cellular processes, environmental information processing, genetic information processing, diseases, metabolism and organismal systems. The results from this study provide a the oretical basis for studying the effects of sbGnRH on the physiology, biochemistry and metabolic enzyme activities of liver in pompano.

Keywords: Trachinotus ovatus; liver; physiology; sbGnRH; transcriptome.

Figure 1

Comparative evaluation of physiological and biochemical indexes in the liver of pompano. (A) protein metabolism; (B) lipid metabolism; (C) energy metabolism; (D) inorganic component; (E) liver function; (F) renal function. Data are expressed as mean ± standard error (SE) (n = 3), and the statistical significance (compared with the control group) was calculated using one-way analysis of variance (ANOVA), followed by Duncan’s post hoc test. * indicate statistical differences at P < 0.05, respectively. ns, not significant (P > 0.05).

Figure 2

Comparative evaluation of related liver enzyme activities between control group and treatment group. (A) digestion; (B) carbohydrate metabolism-related enzymes; (C) antioxidant defense; (D) lipid metabolism; (E) Membrane lipid peroxidation. Data are expressed as mean ± standard error (SEM) (n = 3), and the statistical significance (compared with the control group) was calculated using one-way analysis of variance (ANOVA), followed by Duncan’s post hoc test. * and ** indicate statistical differences at P < 0.05 and P < 0.01, respectively. ns, not significant (P > 0.05).

Figure 3

Comparative distribution of NR homologous species. The Y-coordinate represents species; the X-coordinate represents the number of genes.

Figure 4

Differential gene expression in the liver of pompano between control group and treatment group. (A) Volcano plot of the differences in gene expression. Each dot represents a gene. The green and red dots in the figure represent genes with significant expression differences, green represents down-regulation of gene expression, red represents up-regulation of gene expression, and black dots represent genes with no significant expression differences. (B) Heatmap of the hierarchical cluster of DEGs for illustrating the overall pattern of gene expression among different liver samples.

Figure 5

Enrichment analysis of liver differentially expressed genes (DEGs) gene ontology (GO) between control and sbGnRH injection groups. White bars indicate up-regulated genes; Black bars indicate down-regulated genes; the Y-coordinate represents the number of genes; the X-coordinate represents the name of the pathway.

Figure 6

The top 20 significantly enriched KEGG pathways of differentially expressed genes (DEGs). (A) The pathways and rich factor are shown in the vertical and the horizontal axis, respectively. The dot size indicates the number of genes and the color indicates the q-value. (B) Classification of differentially expressed genes KEGG. The vertical axis is the name of KEGG metabolic pathway, the left part is the specific pathway name, and the right part is the classification category corresponding to each pathway. And the horizontal axis is the Annotated Genes. The number on the column is the number of differentially expressed genes related to this pathway. The same column color represents the same category.

Figure 7

Expression of the gene in the liver of pompano in control and sbGnRH injection groups. (A) Transcriptome data (n=3) and (B) quantitative polymerase chain reaction (qPCR) results (n=3). Data are presented as mean ± standard error (SEM). * and ** indicate statistical differences at P < 0.05 and P < 0.01, respectively. The statistical significance (compared with the control group) was calculated using one-way analysis of variance (ANOVA), followed by Duncan’s post hoc test.