Diverse WGBS profiles of longissimus dorsi muscle in Hainan black goats and hybrid goats

Abstract

Background: Goat products have played a crucial role in meeting the dietary demands of people since the Neolithic era, giving rise to a multitude of goat breeds globally with varying characteristics and meat qualities. The primary objective of this study is to pinpoint the pivotal genes and their functions responsible for regulating muscle fiber growth in the longissimus dorsi muscle (LDM) through DNA methylation modifications in Hainan black goats and hybrid goats.

Methods: Whole-genome bisulfite sequencing (WGBS) was employed to scrutinize the impact of methylation on LDM growth. This was accomplished by comparing methylation differences, gene expression, and their associations with growth-related traits.

Results: In this study, we identified a total of 3,269 genes from differentially methylated regions (DMR), and detected 189 differentially expressed genes (DEGs) through RNA-seq analysis. Hypo DMR genes were primarily enriched in KEGG terms associated with muscle development, such as MAPK and PI3K-Akt signaling pathways. We selected 11 hub genes from the network that intersected the gene sets within DMR and DEGs, and nine genes exhibited significant correlation with one or more of the three LDM growth traits, namely area, height, and weight of loin eye muscle. Particularly, PRKG1 demonstrated a negative correlation with all three traits. The top five most crucial genes played vital roles in muscle fiber growth: FOXO3 safeguarded the myofiber's immune environment, FOXO6 was involved in myotube development and differentiation, and PRKG1 facilitated vasodilatation to release more glucose. This, in turn, accelerated the transfer of glucose from blood vessels to myofibers, regulated by ADCY5 and AKT2, ultimately ensuring glycogen storage and energy provision in muscle fibers.

Conclusion: This study delved into the diverse methylation modifications affecting critical genes, which collectively contribute to the maintenance of glycogen storage around myofibers, ultimately supporting muscle fiber growth.

Keywords: DNA methylation; Glycogen storage; Goats; Muscle fiber; Vasodilatation; Whole-genome bisulfite sequencing.

Fig. 1

Overview of mC contexts in LDM of Hainan black goats and hybrid goats. A Density and (B) levels of mC within three contexts (CG, CHG, CHH) across the genome of LDM tissues in two goat species, with Hainan black goats at the center in A and hybrid goats in B, from innermost to outermost, represent gene density, hybrid mCHH, Hainan mCHH, hybrid mCHG, Hainan mCHG, hybrid mCG, Hainan CG. C Comparison of the average log₂^{(mC number)} in three mC contexts between Hainan black goats and hybrid goats, with the concentric circles indicating log₂^{(mC number)} values of 0, 4, 8, 12, and 16. D Mean mC levels within the three contexts (CG, CHG, and CHH) in various genomic functional regions (promoter, exon, intron, CGI, CGI shore, 3’UTR, 5’UTR, repeat) for LDM tissues in both goat species

Fig. 2

The DMR analysis between Hainan black goats and hybrid goats. A The outermost layer and innermost layers of the circle depict DNA methylation levels for the CG content in hybrid goats and Hainan black goats, respectively. The middle layer represents differences in DNA methylation levels between the two goat species. Each bin in the circos figure was 6,553,600bp. Dark blue bars indicate significantly lower (p < 0.05) DNA methylation levels in hybrid goats compared to Hainan black goats, while red bars signify significantly higher (p < 0.05) DNA methylation levels. B The outer and the inner layers distinguish between hyper and hypo DMR, while the two middle layers show the density of TE and genes. The small red, blue, and purple circles represent CG, CHG, and CHH contexts, respectively. The position of these circles relative to the center indicated the magnitude of the difference in methylation levels (diff.methylation) within DMRs. The size of the circles corresponds to the extent of the difference (areaStat) within DMRs. C Distribution of genes within DMRs across various genomic functional regions. D Comparison of gene functional regions between hyper and hypo DMRs

Fig. 3

Functional analysis of the gene sets within DMRs and DEGs. The A KEGG and B GO enrichment of hyper DMR genes. C KEGG and D GO enrichment of hypo DMR genes. E PPI network of genes intersecting between DMRs and DEGs. The 11 hub genes are highlighted by a red outline

Fig. 4

Functional analysis of gene sets intersecting between DMRs and DEGs. A KEGG enrichment of the intersection gene sets. B Expression patterns of the 11 hub genes from Hainan black goats and hybrid goats. C A depiction of the connections among genes, functional regions, and C contents of hub genes. D Correlations between differential gene expression and the degree of differential methylation levels within hypo DMRs

Fig. 5

Correlations of hub genes with growth traits of LEA. APRKG1 was negatively correlated with the weight, height, and LEA. BNR4A1 was negatively correlated with the height and weight. C-EFOXO3, FOXO6 and GNAO1 were positively correlated with two of the three growth traits. F-IADCY5, AKT2, ZBTB16 were positively correlated with one trait, and EZH2 was negatively correlated with one trait. All hollow circles in A-E indicate Hainan black goats, and solid circles indicate hybrid goats

Fig. 6

Identification and cooperation of hub genes associated with muscle growth. The left side depicts muscle fibers densely interspersed with blood vessels. The right side illustrates a microenvironment encompassing muscle fibers, blood vessels, and phagocytes. The central pink circle, filled with small myofibrils, represents a muscle fiber, surrounded by red and blue blood vessels, and several genes are interconnected around the muscle fiber, jointly contributing to glycogen storage. Red arrows indicate genes with increased expression, while blue arrows indicate genes with decreased expression