Preliminary studies on the molecular mechanism of intramuscular fat deposition in the longest dorsal muscle of sheep

Abstract

Background: Intramuscular fat content is an important index reflecting the quality of mutton, which directly affects the flavor and tenderness of mutton. Livestock and poultry intramuscular fat content is influenced by genetics, nutritional level, and environmental factors. Key regulatory factors play a crucial role in intramuscular fat deposition. However, there is a limited amount of research on the identification and function of key genes involved in intramuscular fat content deposition specifically in sheep.

Results: Histological differences in the longest dorsal muscle of the small-tailed frigid sheep increased in diameter and decreased in several muscle fibers with increasing monthly age; The intramuscular fat content of the longest dorsal muscle of the small-tailed cold sheep varied with age, with a minimum of 1 month of age, a maximum of 6 months of age, and a minimum of 12 months of age. Transcriptomic sequencing and bioinformatics analysis revealed a large number of differential genes in the longest dorsal muscles of little-tailed billy goats of different months of age, which were enriched in multiple GO entries and KEGG pathways. Among them, the pathway associated with intramuscular fat was the AMPK signaling pathway, and the related genes were PPARGC1A and ADIPOQ; Immunohistochemical studies showed that PPARGC1A and ADIPOQ proteins were expressed in connective tissues, cell membranes, and, to a lesser extent, the cytoplasm of the longest dorsal muscle of the little-tailed frigid sheep; Real-time PCR and Western Blot validation showed that PPARGC1A and ADIPOQ were both expressed in the longest dorsal muscle of the little-tailed frigid sheep at different ages, and there were age differences in the amount of expression. The ADIPOQ gene was negatively correlated with the intramuscular fat content of the longest dorsal muscle, and the PPARGC1A gene was positively correlated with the intramuscular fat content of the longest dorsal muscle; As inferred from the above results, the ADIPOQ gene was negatively correlated with the intramuscular fat content of the longest dorsal muscle (r = -0.793, P < 0.05); and the PPARGC1A gene was positively correlated with the intramuscular fat content of the longest dorsal muscle r = 0.923, P < 0.05).

Conclusions: Based on the above results, it can be inferred that the ADIPOQ gene is negatively correlated with the intramuscular fat content of the longest back muscle (r = -0.793, P < 0.05); the PPARGC1A gene is positively correlated with the intramuscular fat content of the longest back muscle (r = 0.923, P < 0.05).

Keywords: Growth and development; Intramuscular fat; Longest dorsal muscle; Small-tailed frigid sheep; Transcriptomics.

Fig. 1

Muscle fiber parameters and intramuscular fat content of the longest dorsal muscle of sheep of different ages. (A) Schematic diagram of the muscle fiber structure of the longest dorsal muscle of sheep, Where “a” is connective tissue, “b” is muscle fiber, and “c” is cell nucleus. (B) HE staining of the longest dorsal muscle of sheep at different months of age. (C) Diameter of muscle fibers of the longest dorsal muscle of sheep at different months of age. (D) Density of muscle fibers in the longest dorsal muscle of sheep at different months of age. (E) Intramuscular fat content of the longest dorsal muscle of sheep at different months of age

Fig. 2

Differentially expressed genes (DEG) between different months of age using 1 month of age as control. (A) Number of differentially differentiated genes at different months of age using 1 month of age as a control; (B) Venn diagram of the comparison of DEGs in the four groups of samples; (C) Volcano plot of DEGs

Fig. 3

GO enrichment and KEGG enrichment of differential genes between different combinations (A) Histogram of GO enrichment analysis of DEGs in different combinations. (B) Scatter plot of KEGG enrichment in different combinations

Fig. 4

The mRNA relative expression level of DEG was verified by qRT-PCR

Fig. 5

Immunohistochemistry and Western blotting test results. (A) Immunohistochemical results of PPARGC1A and ADIPOQ proteins in the intramuscular fat of the longissimus dorsi muscle at different ages. The “a” is nucleus, “b” is cytoplasm and “c” is connective tissue. (B) PPARGC1A and ADIPOQ Western blotting of intramuscular fat in the longest dorsal muscle of sheep of different months of age using 1 month of age as control