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. 2023 Mar 3;14(3):639.
doi: 10.3390/genes14030639.

Genome-Wide Analysis of lncRNA and mRNA Expression in the Uterus of Laying Hens during Aging

Guang Li  1 , Xinyue Yang  1 , Junyou Li  2 , Bingkun Zhang  1
Affiliations

Genome-Wide Analysis of lncRNA and mRNA Expression in the Uterus of Laying Hens during Aging

Guang Li et al. Genes (Basel). .

Abstract

Eggshell plays an essential role in preventing physical damage and microbial invasions. Therefore, the analysis of genetic regulatory mechanisms of eggshell quality deterioration during aging in laying hens is important for the biosecurity and economic performance of poultry egg production worldwide. This study aimed to compare the differences in the expression profiles of long non-coding RNAs (lncRNAs) and mRNAs between old and young laying hens by the method of high-throughput RNA sequencing to identify candidate genes associated with aging in the uterus of laying hens. Overall, we detected 176 and 383 differentially expressed (DE) lncRNAs and mRNAs, respectively. Moreover, functional annotation analysis based on the Gene Ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) databases revealed that DE-lncRNAs and DE-mRNAs were significantly enriched in "phosphate-containing compound metabolic process", "mitochondrial proton-transporting ATP synthase complex", "inorganic anion transport", and other terms related to eggshell calcification and cuticularization. Through integrated analysis, we found that some important genes such as FGF14, COL25A1, GPX8, and GRXCR1 and their corresponding lncRNAs were expressed differentially between two groups, and the results of quantitative real-time polymerase chain reaction (qPCR) among these genes were also in excellent agreement with the sequencing data. In addition, our study found that TCONS_00181492, TCONS_03234147, and TCONS_03123639 in the uterus of laying hens caused deterioration of eggshell quality in the late laying period by up-regulating their corresponding target genes FGF14, COL25A1, and GRXCR1 as well as down-regulating the target gene GPX8 by TCONS_01464392. Our findings will provide a valuable reference for the development of breeding programs aimed at breeding excellent poultry with high eggshell quality or regulating dietary nutrient levels to improve eggshell quality.

Keywords: aging; co-expression network; lncRNA; mRNA; uterus.

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Conflict of interest statement

The authors declare that no conflict of interest exists.

Figures

Figure 1
Figure 1
The features of predicted lncRNAs and mRNAs. (A) Venn diagram of lncRNAs from the Coding Potential Calculator (CPC), the Coding-Non-Coding Index (CNCI), and Protein Families Database (PFAM). (B) Length distribution of lncRNAs and coding transcripts. (C) Exon number distribution of lncRNAs and coding transcripts. (D) Orf length distribution of lncRNAs and coding transcripts.
Figure 2
Figure 2
Analyses of DE-lncRNAs and mRNAs in the eggshell gland. (A) The volcano plot can intuitively see the overall distribution of the differential transcripts, and the threshold value was set to p < 0.05. Blue dots represent that lncRNAs are not significantly differential expressions; Red dots represent relatively high expressions; Green dots represent relatively low expressions. (B) A Heatmap of 176 lncRNA expression profiles showed significant expression differences (91 up-regulated and 85 down-regulated). Data were expressed as FPKM, and the red-to-green color gradient indicates from high expression to low expression. (C) The volcano plot can intuitively see the overall distribution of the differential genes, and the threshold value was set to p < 0.05. Blue dots represent that lncRNAs are not significantly differential expressions; Red dots represent relatively high expressions; Green dots represent relatively low expressions. (D) The Heatmap of 383 mRNAs expression profiles showed significant expression differences (204 up-regulated and 179 down-regulated). Data were expressed as FPKM, and the red-to-green color gradient indicates from high expression to low expression.
Figure 3
Figure 3
LncRNAs-mRNAs co-expression interaction network. DE-lncRNAs (p-adjust < 0.05) and their corresponding differentially expressed cis- and trans-target genes (p-adjust < 0.05) were selected and used to construct a lncRNAs-mRNAs co-expression network. In this network, protein-coding genes are displayed as blue circles, and lncRNA are displayed as pink diamonds. Solid lines mean the interactions between DE-lncRNAs and their corresponding cis-target genes, whereas the dashed lines mean interactions between DE-lncRNAs and their corresponding trans-target genes.
Figure 4
Figure 4
Validation of 4 DE-lncRNAs and their target genes by qPCR. The qPCR results of the DE-lncRNAs and DEGs were compared with their RNA-Seq, respectively. Red: represents qPCR, and Blue: represents RNA-seq.
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This study was supported by the National Key R&D Program of China (2022YFE0111100) and by the 2115 Talent Development Program of China Agricultural University.

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