Comparative proteomic analysis of the ovarian fluid and eggs of Siberian sturgeon

Abstract

Background: Sturgeon species are living fossils that exhibit unique reproductive characteristics, and elucidation of the molecular processes governing the formation and quality of sturgeon eggs is crucial. However, comprehensive data on the protein composition of sturgeon ovarian fluid (OF) and eggs and their functional significance are lacking. To address this knowledge gap, the aim of the present study was to conduct a comprehensive comparative proteomic analysis of Siberian sturgeon OF and eggs using liquid chromatography-mass spectrometry (LC-MS/MS).

Results: A total of 617 proteins were identified in OF, and 565 proteins were identified in eggs. A total of 772 proteins showed differential abundance. Among the differentially abundant proteins, 365 were more abundant in OFs, while 407 were more abundant in eggs. We identified 339 proteins unique to OFs and 287 proteins specific to eggs, and further investigated the top 10 most abundant proteins in each. The functional annotation of the OF proteins highlighted their predominant association with immune system processes, including the complement and coagulation cascade, neutrophil and leukocyte-mediated immunity, cholesterol metabolism, and regulation of the actin cytoskeleton. Analysis of egg proteins revealed enrichment in metabolic pathways, such as oxidative phosphorylation and fatty acid metabolism, and protein ubiquitination and translation. OF-specific proteins included extracellular matrix and secretory vesicles, and eggs were enriched in proteins localized to mitochondria and ribosome components.

Conclusions: This study presents the first comprehensive characterization of the protein composition of sturgeon OF and eggs and elucidates their distinct functional roles. These findings advance our understanding of sturgeon reproduction, OF-egg signaling and the origin of OF proteins. The mass spectrometry proteomics data have been deposited in the ProteomeXchange Consortium with the dataset identifier PXD044168 to ensure accessibility for further research.

Keywords: Acipenser baerii; Eggs; Mass spectrometry; Ovarian fluid; Proteome.

Fig. 1

GO biological process enrichment analysis of total identified proteins in the ovarian fluid (OF) and eggs of Siberian sturgeon. Most OF proteins were associated with exocytosis and secretion and immune system processes, but the top-ranked biological processes were cellular metabolic and catabolic processes in eggs

Fig. 2

Functional analysis of proteins unique to ovarian fluid (OF) and eggs. Venn diagram representing the overlap of all identified proteins in the OF and eggs of Siberian sturgeon (A). GO biological processes and cellular components; KEGG pathway analysis of the OF (B) and egg (C) proteins. Pathways with many shared genes are clustered together. Larger dots indicate more significant P values

Fig. 3

Results of the Search Tool for the Retrieval of Interacting Genes/Proteins (v 11.0) analysis showing the protein association network of proteins specific to ovarian fluid assigned to immune system processes with a high confidence score. The nodes correspond to the proteins, and the edges represent the interactions (thick lines indicate a high score > 0.9; thin lines indicate a medium score > 0.7). Model statistics are presented on the left. An explanation of the edge colors is provided above the figure

Fig. 4

Results of the Search Tool for the Retrieval of Interacting Genes/Proteins (v 11.0) analysis showing the protein association network of proteins specific to eggs assigned to metabolic processes with a high confidence score. The nodes correspond to the proteins, and the edges represent the interactions (thick lines indicate a high score > 0.9; thin lines indicate a medium score > 0.7). Model statistics are presented on the left. An explanation of the edge colors is provided above the figure

Fig. 5

Validation of proteins specific to ovarian fluid (OF) and eggs, such as albumin (ALB), fibrinogen (FGB), fibronectin-1 (FN1), and vitellogenin 2 (VTG2), using Western blot analysis (A). The full-length blots are presented in Supplementary Fig. S2. A quantitative comparison of VTG2 in the OF and eggs (B). The data are presented as the means ± S.D. (n = 5 in OF and eggs). Statistical analysis was performed using Student’s t test; ****p ≤ 0.0001. The intensity of the protein bands on the TGX Stain-Free gels was analyzed using Image Lab 6 software (Bio-Rad)

Fig. 6

Proteins identified in Siberian sturgeon eggs that participate in mitochondrial oxidative phosphorylation complexes I–V (A) and fatty acid -oxidation I (B). The scheme was prepared using BioRender software. Descriptions of the gene/protein symbols are presented in Supplementary Table S2

Fig. 7

Scheme presenting an overall summary of the unique roles of the identified proteins in the ovarian fluid (OF) and eggs of Siberian sturgeon. OF proteins were associated with the inflammatory cascade, modulation of the extracellular matrix and lipid metabolism, and proteins in the eggs were primarily involved in embryo development, fertilization, translation, ubiquitin - proteasome degradation and energy metabolism. Descriptions of the gene/protein symbols are presented in Supplementary Table S2