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. 2022 Apr 5:9:854503.
doi: 10.3389/fvets.2022.854503. eCollection 2022.

Comparison of miRNA and mRNA Expression in Sika Deer Testes With Age

Boyin Jia  1   2 , Linlin Zhang  1 , Fuquan Ma  1 , Xue Wang  1 , Jianming Li  2   3 , Naichao Diao  1   2 , Xue Leng  2   3 , Kun Shi  2   3 , Fanli Zeng  2   3 , Ying Zong  2   3 , Fei Liu  1   2 , Qinglong Gong  1   2 , Ruopeng Cai  1   2 , Fuhe Yang  4 , Rui Du  2   3 , Zhiguang Chang  5
Affiliations

Comparison of miRNA and mRNA Expression in Sika Deer Testes With Age

Boyin Jia et al. Front Vet Sci. .

Abstract

To elucidate the complex physiological process of testis development and spermatogenesis in Sika deer, this study evaluated the changes of miRNA and mRNA profiles in the four developmental stages of testis in the juvenile (1-year-old), adolescence (3-year-old), adult (5-year-old), and aged (10-year-old) stages. The results showed that a total of 198 mature, 66 novel miRNAs, and 23,558 differentially expressed (DE) unigenes were obtained; 14,918 (8,413 up and 6,505 down), 4,988 (2,453 up and 2,535 down), and 5,681 (2,929 up and 2,752 down) DE unigenes, as well as 88 (43 up and 45 down), 102 (44 up and 58 down), and 54 (18 up and 36 down) DE miRNAs were identified in 3- vs. 1-, 5- vs. 3-, and 10- vs. 5-year-old testes, respectively. By integrating miRNA and mRNA expression profiles, we predicted 10,790 mRNA-mRNA and 69,883 miRNA-mRNA interaction sites. The target genes were enriched by GO and KEGG pathways to obtain DE mRNA (IGF1R, ALKBH5, Piwil, HIF1A, BRDT, etc.) and DE miRNA (miR-140, miR-145, miR-7, miR-26a, etc.), which play an important role in testis development and spermatogenesis. The data show that DE miRNAs could regulate testis developmental and spermatogenesis through signaling pathways, including the MAPK signaling pathway, p53 signaling pathway, PI3K-Akt signaling pathway, Hippo signaling pathway, etc. miR-140 was confirmed to directly target mutant IGF1R-3'UTR by the Luciferase reporter assays. This study provides a useful resource for future studies on the role of miRNA regulation in testis development and spermatogenesis.

Keywords: mRNA; miRNA; sika deer; spermatogenesis; testis development.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Statistics for DE unigenes and miRNAs in each comparable group. (A) q value < 0.005 and |log2 (fold change)| >1 were used as thresholds of significance for DE unigenes. (B) q value < 0.01 and |log2 (fold change)| >1 were used as thresholds of significance for DE miRNAs.
Figure 2
Figure 2
Interaction networks of top DE unigenes in each comparable group. (A) Tst_2 vs. Tst_1. (B) Tst_3 vs. Tst_2. (C) Tst_4 vs. Tst_3. The up-regulated unigenes were displayed as red circles, and the down-regulated unigenes were displayed as green circles.
Figure 3
Figure 3
Hierarchical clustering of DE miRNAs.
Figure 4
Figure 4
miRNA-mRNA interaction network related to testis development of Sika Deer. (A) Up-regulated miRNAs and down-regulated target genes related to testis development. (B) Down-regulated miRNAs and up-regulated target genes related to testis development. The up-regulated miRNAs or genes were displayed as red circles, and the down-regulated miRNAs or genes were displayed as green circles.
Figure 5
Figure 5
miRNA-mRNA interaction network related to spermatogenesis of Sika Deer. (A) Up-regulated miRNAs and down-regulated target genes related to spermatogenesis. (B) Down-regulated miRNAs and up-regulated target genes related to spermatogenesis. The up-regulated miRNAs or genes were displayed as red circle, and the down-regulated miRNAs or genes were displayed as green circles.
Figure 6
Figure 6
Q-PCR validation of mRNA-seq data.
Figure 7
Figure 7
Q-PCR validation of miRNA-seq data.
Figure 8
Figure 8
Detection of interactions between miR-140 and IGF1R by dual luciferase reporter system. (A) Binding site sequence of miR-140 and target gene IGF1R. (B) miR-140 mimic was cotransfected with IGF1R 3 'UTR WT or IGF1R 3 'UTR MUT to detect the luciferase activity for 48 h. *p < 0.05.
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