Min pig skeletal muscle response to cold stress

Abstract

The increased sensitivity of pigs to ambient temperature is due to today's intensive farming. Frequent climate disasters increase the pressure on healthy pig farming. Min pigs are an indigenous pig breed in China with desirable cold resistance characteristics, and hence are ideal for obtaining cold-resistant pig breeds. Therefore, it is important to discover the molecular mechanisms that are activated in response to cold stress in the Min pig. Here, we conducted a transcriptomic analysis of the skeletal muscle of Min pigs under chronic low-temperature acclimation (group A) and acute short cold stress (group B). Cold exposure caused more genes to be upregulated. Totals of 125 and 96 differentially expressed genes (DEGs) were generated from groups A and B. Sixteen common upregulated DEGs were screened; these were concentrated in oxidative stress (SRXN1, MAFF), immune and inflammatory responses (ITPKC, AREG, MMP25, FOSL1), the nervous system (RETREG1, GADD45A, RCAN1), lipid metabolism (LRP11, LIPG, ITGA5, AMPD2), solute transport (SLC19A2, SLC28A1, SLCO4A1), and fertility (HBEGF). There were 102 and 73 genes that were specifically differentially expressed in groups A and B, respectively. The altered mRNAs were enriched in immune, endocrine, and cancer pathways. There were 186 and 91 differentially expressed lncRNAs generated from groups A and B. Analysis of the target genes suggested that they may be involved in regulating the MAPK signaling pathway for resistance to cold. The results of this study provide a comprehensive overview of cold exposure-induced transcriptional patterns in skeletal muscle of the Min pig. These results can guide future molecular studies of cold stress response in pigs for improving cold tolerance as a goal in breeding programs.

Fig 1. DEGs in the skeletal muscle of Min pig after cold stress.

A. Volcano plot for chronic low-temperature acclimation group (A) after cold stress. Red points represent the upregulated genes; green points represent the downregulated genes (|log₂FC|>0.59 and p< 0.05), and blue points represent the not significantly DEGs (p> 0.05). B. Volcano plot for acute short cold stress group (B) after cold stress. The means of color points are similar to A. C. Total DEGs in the groups A and B after cold stress. D. Upregulated DEGs in the groups A and B after cold stress. E. Downregulated DEGs in the groups A and B after cold stress.

Fig 2. Pathway analysis of DEGs.

A. KEGG enrichment analysis of upregulated genes for chronic low-temperature acclimation group (-log₁₀ (p-value)>1.3). B. KEGG enrichment analysis of downregulated genes for chronic low-temperature acclimation group (-log₁₀ (p-value)>1.3). C. KEGG enrichment analysis of upregulated genes for acute short cold stress group (-log₁₀ (p-value)>1.3). D. KEGG enrichment analysis of downregulated genes for acute short cold stress group (-log₁₀ (p-value)>1.3).

Fig 3. Differentially expressed lncRNAs in the skeletal muscle of the Min pig under cold stress.

A. Total differentially expressed lncRNAs in the chronic low-temperature acclimation group and acute short cold stress group (log₂FC >1 and q < 0.05). B. Upregulated differentially expressed lncRNAs in the chronic low-temperature acclimation group and acute short cold stress group. C. Downregulated differentially expressed lncRNAs in the chronic low-temperature acclimation group and acute short cold stress group. D. The distribution of lncRNAs on pig chromosomes for chronic low-temperature acclimation group. The maximum circle represents the chromosome length of the pig genome; the second circle represents all lncRNAs detected in this experiment, and the smallest circle represents the significantly differentially expressed lncRNAs (|log ₂FC|>1 and q<0.05). E. The distribution of lncRNAs on pig chromosomes for acute short cold stress group. The means of every circle is same to D. F. The proportions of three categories lncRNAs for chronic low-temperature acclimation group and acute short cold stress group.

Fig 4. lncRNA-mRNA-network analysis.

A. Blue nodes represent upregulated mRNAs; red nodes represent upregulated lncRNAs, and green nodes represent downregulated lncRNAs. B, C, and D illustrate GO analyses of target mRNAs of differentially expressed lncRNAs of the chronic low-temperature acclimation group. E, F and G illustrate GO analyses of target mRNAs of differentially expressed lncRNAs of the acute short cold stress group.

Fig 5. The differential expression of lncRNAs and mRNAs validated by quantitative real-time PCR (qRT-PCR).

A. The qRT-PCR and RNA-seq results of chronic low-temperature acclimation group. B. The qRT-PCR and RNA-seq results of acute short cold stress group.