Comparative analysis of differentially abundant proteins between high and low intramuscular fat content groups in donkeys

Abstract

Intramuscular fat (IMF) is an important regulator that determines meat quality, and its content is closely related to flavor, tenderness, and juiciness. Many studies have used quantitative proteomic analysis to identify proteins associated with meat quality traits in livestock, however, the potential candidate proteins that influence IMF in donkey muscle are not fully understood. In this study, we performed quantitative proteomic analysis, with tandem-mass-tagged (TMT) labeling, with samples from the longissimus dorsi (LD) muscle of the donkey. A total of 585,555 spectra were identified from the six muscle samples used in this study. In total, 20,583 peptides were detected, including 15,279 unique peptides, and 2,540 proteins were identified. We analyzed differentially abundant proteins (DAPs) between LD muscles of donkeys with high (H) and low (L) IMF content. We identified 30 DAPs between the H and L IMF content groups, of which 17 were upregulated and 13 downregulated in the H IMF group. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analysis of these DAPs revealed many GO terms (e.g., bone morphogenetic protein (BMP) receptor binding) and pathways (e.g., Wnt signaling pathway and Hippo signaling pathway) involved in lipid metabolism and adipogenesis. The construction of protein-protein interaction networks identified 16 DAPs involved in these networks. Our data provide a basis for future investigations into candidate proteins involved in IMF deposition and potential new approaches to improve meat quality in the donkey.

Keywords: IMF; differentially abundant proteins; donkey; functional analysis; proteomics.

Figure 1

Schematic protocol for the identification of longissimus dorsi muscle proteins from tissue samples with high (H) and low (L) intramuscular fat (IMF) content using tandem-mass-tagged (TMT)-labeled quantitative proteomics. The H and L IMF samples were in triplicate, and each sample used a different TMT label.

Figure 2

Characterization of proteins from six longissimus dorsi muscle samples. (A) Overview information of identified proteins in this study. (B) Distribution of the lengths of the identified peptides. (C) Distribution of the numbers of identified proteins containing different numbers of peptides. (D) Distribution of the molecular weights of the identified proteins.

Figure 3

Analysis of differentially abundant proteins (DAPs) between high and low intramuscular fat (IMF) groups. (A) Volcano plot showing DAPs between high and low IMF groups. Red, gray, and blue dots represent upregulated, unchanged, and downregulated proteins, respectively. (B) Heatmap showing the abundance patterns of the DAPs between the three high and three low IMF samples.

Figure 4

Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analysis of differentially abundant proteins (DAPs). (A) Top 30 significantly enriched GO terms. The X-axis represents the –log10 (p value) value of the GO terms and the Y-axis represents the GO term name. (B) Significantly enriched pathways of DAPs. The X-axis represents the rich factor and the Y-axis represents the pathway. The size and color of the bubbles represent the number of proteins enriched in the pathway and enrichment significance, respectively.

Figure 5

Protein–protein interactions analysis for differentially abundant proteins (DAPs). The red nodes represent DAPs, while the green nodes represent other proteins that interact with DAPs. The edge represents interactions between proteins.