Transcriptome analysis of mRNA and microRNAs in intramuscular fat tissues of castrated and intact male Chinese Qinchuan cattle.

Ying-Ying Zhang,Yang Yang,Dai Chen,Hong-Bao Wang,Hong-Fang Guo,Hong-Cheng Wang,Lin-Sen Zan,Song Zhang,Ya-Ning Wang,Jie-Yun Hong

doi:10.1371/journal.pone.0185961

Abstract

Intramuscular fat (IMF) is known to enhance beef palatability and can be markedly increased by castration. However, there is little understanding of the molecular mechanism underlying the IMF deposition after castration of beef cattle. We hypothesize that genetic regulators function differently in IMF from bulls and steers. Therefore, after detecting serum testosterone and lipid parameter, as well as the contents of IMF at 6, 12, 18 and 24 months, we have investigated differentially expressed (DE) microRNAs (miRNAs) and mRNAs in IMF of bulls and steers at 24 months of age in Qinchuan cattle using next-generation sequencing, and then explored the possible biopathways regulating IMF deposition. Serum testosterone levels were significantly decreased in steers, whereas IMF content, serum total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and triglycerides (TGs) were markedly increased in steers. Comparing the results of steers and bulls, 580 upregulated genes and 1,120 downregulated genes in IMF tissues were identified as DE genes correlated with IMF deposition. The upregulated genes were mainly associated with lipid metabolism, lipogenesis and fatty acid transportation signalling pathways, and the downregulated genes were correlated with immune response and intracellular signal transduction. Concurrently, the DE miRNAs—important players in adipose tissue accumulation induced by castration—were also examined in IMF tissues; 52 DE miRNAs were identified. The expression profiles of selected genes and miRNAs were also confirmed by quantitative real-time PCR (qRT-PCR) assays. Using integrated analysis, we constructed the microRNA-target regulatory network which was supported by target validation using the dual luciferase reporter system. Moreover, Ingenuity Pathway Analysis (IPA) software was used to construct a molecular interaction network that could be involved in regulating IMF after castration. The detected molecular network is closely associated with lipid metabolism and adipocyte differentiation, which is supported by functional identification results of bta-let-7i on bovine preadipocytes. These results provided valuable insights into the molecular mechanisms of the IMF phenotype differences between steers and bulls.

Highlights

The amount of intramuscular fat (IMF) in a cross-section of muscle tissue is highly correlated with meat palatability, affecting the juiciness, tenderness and flavor
We found that the expression of mitogen-activated protein kinase kinase kinase 1 (MAP3K1), carnitine palmitoyltransferase 1A (CPT1A) and protein kinase cAMPdependent catalytic beta (PRKACB) was downregulated in steers; in contrast, lipoprotein lipase (LPL), fatty acid synthase (FASN), diacylglycerol O-acyltransferase 2 (DGAT2), 1-acyl-sn-glycerol-3-phosphate acyltransferase (AGPAT2) and Cyclic AMP-responsive element-binding protein 3-like protein 1 (CREB3L1) were significantly upregulated in steers. quantitative realtime PCR (qRT-PCR) analysis largely confirmed the RNA-seq data
We found that several genes involved in mitogen-activated protein kinases (MAPK) signaling pathway, such as MAP3K1, an MAPK kinase 4 (MKK4) activator, related C3 botulinum toxin substrate 2 (RAC2), a MAP3K1 activator, and JUN, a c-Jun NH 2-terminal kinase (JNK) activator, were all downregulated in the IMF of steers [59,60]

Summary

Introduction

The amount of intramuscular fat (IMF) in a cross-section of muscle tissue is highly correlated with meat palatability, affecting the juiciness, tenderness and flavor. Efforts to improve production efficiency and select the animals with high-lean growth rate have negatively influenced the deposition of IMF [1,2]. Enhancing intramuscular adipose deposition while maintaining stable levels of lipid deposition in other depots have become an important issue in beef industry. It is essential to advance our understanding of the molecular mechanisms that affect IMF deposition so that genomic selection can be used to produce high-grade beef with consistent quality. Steers and bulls are good animal models for the comparative study of IMF differences. The mechanisms underlying differences in the deposition of IMF after castration are poorly understood

Methods

Results

Discussion

Conclusion