| SIRT family is a NAD+ dependent class III deacetylase family, which is involved in modifying histone or non-histone proteins. In addition to having deacetylase, some members also have ADP-ribosylase and other activities, which play an important role in regulating energy metabolism and resisting oxidative stress. The SIRT family exists widely in prokaryotes and eukaryotes, and is mainly divided into 5 classes. SIRT1-SIRT3 is class Ⅰ, SIRT4 is class Ⅱ, SIRT5 is class Ⅲ, SIRT6 and SIRT7 are class Ⅳ, and class U exists only in the SIRT family from archaea to bacteria. The number and distribution of SIRT genes were different among different organisms. All members of the SIRT family have been shown to be expressed in mammalian ovaries and are widely involved in regulating ovarian development, including meiosis regulation, energy metabolism, mitochondrial quality control, maintenance of redox homeostasis, and hormone secretion. In this study, bioinformatics methods were used to systematically analyze the chromosome distribution, gene structure, amino acid sequence, protein motifs and conserved domains, physical and chemical properties, subcellular localization, secondary and tertiary structures, phylogenetic relationships, and protein interaction networks of zebrafish SIRT family genes, and to explore their expression changes at different developmental stages of follicles. The results showed that 8 SIRT genes were distributed on 8 chromosomes of Zebrafish, the sequence length was different, the longest was SIRT6(140265bp), the shortest was SIRT4(7101bp), and the coding regions ranged from 3 to 14. The amino acid sequence similarity of zebrafish SIRT protein was low, and the 10 most conserved motifs of zebrafish SIRT protein were predicted. The adjacent homologous genes in the same branch had nearly the same motif composition, and the number of motifs was different among branches. Among the 10 motifs, motif 2, motif 3 and motif 5 were found in all zebrafish SIRT amino acid sequences, indicating that these protein motifs are highly conserved during development. Conserved domain analysis showed that all SIRT1-7 proteins contained the Sir2 domain. The analysis of physical and chemical properties of proteins showed that the largest molecular weight was SIRT1, encoding 710 amino acids, while the smallest molecular weight was SIRT5, encoding 305 amino acids. The isoelectric points ranged from 4.88 to 9.60, and all of them were hydrophilic proteins. Except SIRT3, the rest were unstable proteins. Subcellular localization prediction showed that SIRT1, SIRT4, SIRT5 and SIRT7 were located in cytoplasm/nucleus, SIRT3.2 in cytoplasm, SIRT6 in nucleus, SIRT2 in cytoskeleton, and SIRT3 in mitochondria. The results of secondary structure analysis showed that the SIRT family proteins had similar secondary structure, and α-helix and random curling were the main components of the protein secondary structure. The tertiary structure prediction showed that the SIRT protein family had zinc finger structure and Rossmann fold structure. Phylogenetic analysis showed that the fish SIRT family could be divided into three branches. The first branch consisted of three subbranches, in which SIRT1 and SIRT2 were isolated and SIRT3 and SIRT3.2 were clustered into one branch. The second largest branch consisted of SIRT4 and SIRT5, which were clustered separately into one branch. The third branch consists of SIRT6 and SIRT7, each of which is a separate branch. The 8 SIRT proteins of Zebrafish had low homology and were distributed far in the evolutionary tree. Compared with other species, zebrafish SIRT1 is closely related to rainbow trout SIRT1, while other family members are closely related to goldfish and electric eel SIRT. Collinearity analysis showed that four family members (SIRT1, SIRT2, SIRT3.2, SIRT4) were collinearity between the blue killifish and zebrafish, while the other family members except SIRT6 were collinearity between the goldfish and zebrafish. In addition, zebrafish SIRT4 and SIRT5 were collinearity with two genes of goldfish, respectively. PPI prediction showed that SIRT protein interacts with ESR1, FOXOs, SOD, HSPs, etc. Real-time fluorescence quantitative PCR analysis showed that SIRT1 and SIRT2 were mainly expressed in MV stage during follicular development. SIRT3 and SIRT4 were mainly expressed at FG stage. SIRT3.2, SIRT5, SIRT6 and SIRT7 were mainly expressed in GVBD stage. In summary, this study used bioinformatics methods for the first time to analyze chromosome localization, gene structure analysis, amino acid sequence analysis, physical and chemical properties analysis, subcellular localization prediction, phylogenetic analysis, PPI network prediction and follicle expression analysis at different developmental stages of zebrafish SIRT gene family. The results showed that the gene structure and amino acid sequence of 8 members of the Zebrafish SIRT family were different, but all had Sir2 conserved domain and similar protein structure. Phylogenetic analysis suggests that there may be replication or fusion events among SIRT gene family members in different species. SIRT is expressed in zebrafish follicles at different developmental stages with different expression patterns, suggesting that SIRT plays an important role in the regulation of follicle development, providing a reference for further functional studies and the study of complex molecular regulatory network of fish follicle development. |
