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Integrated miRNA-mRNA analysis reveals regulatory pathways underlying the curly fleece trait in Chinese tan sheep.

ABSTRACT: Tan sheep is an indigenous Chinese breed well known for its beautiful curly fleece. One prominent breed characteristic of this sheep breed is that the degree of curliness differs markedly between lambs and adults, but the molecular mechanisms regulating the shift are still not well understood. In this study, we identified 49 differentially expressed (DE) microRNAs (miRNAs) between Tan sheep at the two stages through miRNA-seq, and combined the data with that in our earlier Suppression Subtractive Hybridization cDNA (SSH) library study to elucidate the mechanisms underlying curly fleece formation.Thirty-six potential miRNA-mRNA target pairs were identified using computational methods, including 25 DE miRNAs and 10 DE genes involved in the MAPK signaling pathway, steroid biosynthesis and metabolic pathways. With the differential expressions between lambs and adults confirmed by qRT-PCR, some miRNAs were already annotated in the genome, but some were novel miRNAs. Inhibition of KRT83 expression by miR-432 was confirmed by both gene knockdown with siRNA and overexpression, which was consistent with the miRNAs and targets prediction results.Our study represents the comprehensive analysis of mRNA and miRNA in Tan sheep and offers detailed insight into the development of curly fleece as well as the potential mechanisms controlling curly hair formation in humans.

SUBMITTER: Liu Y

PROVIDER: S-EPMC5948824 | biostudies-other | 2018 May

REPOSITORIES: biostudies-other

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Integrated miRNA-mRNA analysis reveals regulatory pathways underlying the curly fleece trait in Chinese tan sheep.

Liu Yufang Y Zhang Jibin J Xu Qiao Q Kang Xiaolong X Wang Kejun K Wu Keliang K Fang Meiying M

BMC genomics 20180511 1

<h4>Background</h4>Tan sheep is an indigenous Chinese breed well known for its beautiful curly fleece. One prominent breed characteristic of this sheep breed is that the degree of curliness differs markedly between lambs and adults, but the molecular mechanisms regulating the shift are still not well understood. In this study, we identified 49 differentially expressed (DE) microRNAs (miRNAs) between Tan sheep at the two stages through miRNA-seq, and combined the data with that in our earlier Sup ...[more]

PMID: 29751742

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Project description:Small Tail Han Sheep is a highly valued local breed in China because of their precocity, perennial estrus, and high fecundity. The average annual lambing rate of ewes is as high as 180-270%, the semen of ram has characteristics of high yield, high density, and good motility. To reveal the key miRNAs and miRNA-targets underlying testis development and spermatogenesis in male Small Tail Han Sheep, integrated analysis of miRNA and mRNA expression profiles in 2-, 6-, and 12-month-old testes was performed by RNA-seq technology and bioinformatics methods. The results showed that total of 153 known sheep miRNAs and 2712 novel miRNAs were obtained in 2-,6 - and 12-month-old Small Tail Han Sheep testes; 5, 1, and 4 differentially expressed (DE) known sheep miRNAs, and 132, 105, and 24 DE novel miRNAs were identified in 2- vs. 6-, 6- vs. 12-, and 2- vs. 12-month-old testes, respectively. We combined miRNA results of this study and the mRNA results obtained in our previous study to predict the target mRNAs of DE known sheep miRNAs; 131, 10, and 15 target mRNAs of DE known sheep miRNAs and 76, 1, and 11 DE miRNA-targets were identified in the three groups, respectively. GO and KEGG analyses showed that: in 2- vs. 6-month-olds, the target genes of DE known sheep miRNAs were involved in 100 biological processes and 11 signaling pathways; in 6- vs. 12-month-olds, the target genes of DE known sheep miRNAs were involved in 4 biological processes; and in 2- vs. 12-month-olds, the target genes of DE known sheep miRNAs were involved in 17 biological processes and 4 signaling pathways. Three miR-target regulatory networks were constructed based on these DE miRNA-targets. The key miRNA-Targets involved in testis development and spermatogenesis were screened. 6 known sheep miRNAs and 6 novel miRNAs were selected to validate the accuracy of miRNA sequencing data by qRT-PCR. The binding sites of oar-miR-379-5p with WNT8A was validated by a dual luciferase reporter gene detection system.

Project description:Tan sheep (Ovis aries), a Chinese indigenous breed, has special curly fleece after birth, especially at one month old. However, this unique phenotype disappears gradually with age and the underlying reasons of trait evolvement are still unknown. In this study, skin transcriptome data was used to study this issue. In total 51,215 transcripts including described transcripts and transfrags were identified. Pathway analysis of the top 100 most highly expressed transcripts, which included TCHH and keratin gene family members, such as KRT25, KRT5, KRT71, KRT14 and others, showed pathways known to be relevant to hair/fleece development and function. Six hundred differentially expressed (DE) transcripts were detected at two different physiological ages (one-month-old with curly fleece and 48-month-old without curly fleece) and were categorized into three major functional groups: cellular component, molecular function, and biological process. The top six functional categories included cell, cell part, cellular process, binding, intracellular, metabolic process. The detected differentially expressed genes were particularly involved in signal, signal peptide, disulfide bond, glycoprotein and secreted terms, respectively. Further splicing isoform analysis showed that the metallothionein 3 isoform was up-regulated in Tan lamb skin, indicating that it may be related to the conformation of curly fleece in Chinese Tan lamb. The hair-related important differentially expressed genes (SPINK4, FGF21, ESRα, EphA3, NTNG1 and GPR110) were confirmed by qPCR analysis. We deduced that the differences existed in expressed transcripts, splice isoforms and GO categories between the two different physiological stages, which might constitute the major reasons for explaining the trait evolvement of curly fleece in Chinese Tan sheep. This study provides some clues for elucidating the molecular mechanism of fleece change with age in Chinese Tan sheep, as well as supplying some potential values for understanding human hair disorder and texture changes.

Project description:BackgroundThe adrenal gland participates in the process of sheep reproduction. MicroRNAs (miRNAs), endogenous small noncoding RNAs, regulate gene expression at the posttranscriptional level. However, the miRNA-mRNA network profile of adrenal glands relating to reproduction in sheep is still not well-studied. As sheep with FecBBB genotype show higher lambing number compare with the sheep with FecB++ genotype. This research aims to compare gene expression by small RNA-seq in adrenal tissues at follicular (F) and luteal (L) phases in FecBBB (MM) and FecB++ (ww) sheep. After analysis of gene expression, significant differentially expressed microRNAs (DEMs) and corresponding target genes were identified.ResultsA total of 180 miRNAs were found in this study, of which 19 DEMs were expressed in the four comparison groups (MM_F_A vs. MM_L_A, MM_F_A vs. ww_F_A, MM_L_A vs. ww_L_A, ww_F_A vs. ww_L_A). Subsequently, 354 target genes of 19 DEMs were predicted by integrated analysis. Cluster analysis was performed by K_means_cluster, and the expression patterns of these DEMs were separated into four subclusters. Functional analysis of target genes was performed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). The results indicated that the target genes were involved mainly in the Notch signaling pathway, signal transduction, cell communication, innate immune response and amino acid metabolism. Specifically, the Notch signaling pathway, biosynthetic process and metabolic process of pyrimidine nucleotide and amino acid metabolism appear to play key regulatory roles in the sheep fertility trait. Furthermore, miRNA-mRNA interaction networks were constructed by differentially expressed genes combined with our previous study of transcriptome data. The results showed that several key genes, including TDRD3, ANAPC7, CCNL2, BRD2 and MUT, were related to the transformation from the follicular phase to the luteal phase. PLAC8L1, NFAT5, DDX24 and MBD1 were related to the high fecundity of small tail Han sheep.ConclusionsIn this study, the miRNA transcriptome profile was identified, and miRNA-mRNA interaction networks were constructed in adrenal gland tissue of small tail Han sheep, the interaction between miR-370-3p and its targets were considered to play a major role in the reproduction regulation process. The results enriched the number of known miRNAs in adrenal glands and provided novel ideas and further information to demonstrate posttranscriptional regulation mechanisms at follicular and luteal phases in different genotypes of small tail Han sheep.

Project description:BACKGROUND:Yam tuber is a storage organ, derived from the modified stem. Tuber expansion is a complex process, and depends on the expressions of genes that can be influenced by environmental and endogenous factors. However, little is known about the regulatory mechanism of tuber expansion. In order to identify the genes and miRNAs involved in tuber expansion, we examined the mRNAs and small RNAs in Dioscorea opposita (Chinese yam) cv. Guihuai 16 tuber during its initiation and expansion stages. RESULTS:A total of 14,238 differentially expressed genes in yam tuber at its expansion stage were identified by using RNA sequencing technology. Among them, 5723 genes were up-regulated, and 8515 genes were down-regulated. Functional analysis revealed the coordination of tuber plant involved in processes of cell events, metabolism, biosynthesis, and signal transduction pathways at transcriptional level, suggesting that these differentially expressed genes are somehow involved in response to tuber expansion, including CDPK, CaM, CDL, SAUR, DELLA, SuSy, and expansin. In addition, 541 transcription factor genes showed differential expression during the expansion stage at transcriptional level. MADS, bHLH, and GRAS were involved in cell differentiation, division, and expansion, which may relate to tuber expansion. Noteworthy, data analysis revealed that 22 known tuber miRNAs belong to 10 miRNA families, and 50 novel miRNAs were identified. The integrated analysis of miRNA-mRNA showed that 4 known miRNAs and 11 genes formed 14 miRNA-target mRNA pairs were co-expressed in expansion stage. miRNA160, miRNA396, miRNA535 and miRNA5021 may be involved in complex network to regulate cell division and differentiation in yam during its expansion stage. CONCLUSION:The mRNA and miRNA datasets presented here identified a subset of candidate genes and miRNAs that are putatively associated with tuber expansion in yam, a hypothetical model of genetic regulatory network associated with tuber expansion in yam was put forward, which may provide a foundation for molecular regulatory mechanism researching on tuber expansion in Dioscorea species.

Project description:Background Tan sheep, an important local sheep breed in China, is famous for their fur quality. One-month-old Tan sheep have white, curly hair with beautiful flower spikes, commonly known as “nine bends”, which has high economic value. However, the “nine bends” characteristic gradually disappears with age; consequently, the economic value of the Tan sheep decreases. Age-related changes in DNA methylation have been reported and may be responsible for age-induced changes in gene expression. Until now, no genome-wide surveys have been conducted to identify potential DNA methylation sites involved in different sheep growth stages. In this study we investigated the dynamic changes of genome-wide DNA methylation profiles in Tan sheep using DNA from skin and deep whole-genome bisulfite sequencing, and compared the DNA methylation levels at three different growth stages: 1, 24, and 48 months old (mon1, mon24, and mon48, respectively). Results In this study, 11 skin samples from three growth stages (four for mon1, four for mon24, and three for mon48) were used for DNA methylation analysis and gene expression profiling. There were 52, 288 and 236 differentially methylated genes (DMGs) identified between mon1 and mon24, mon1 and mon48, and mon24 and mon48, respectively. Of the differentially methylated regions, 1.11%, 7.61%, and 7.65% were in the promoter in mon1 vs. mon24, mon24 vs. mon48, and mon1 vs. mon48, respectively. DMGs were enriched in the MAPK and WNT signaling pathways, which are related to age growth and hair follicle morphogenesis processes. There were 51 DMGs associated with age growth and curly fleece formation. Four DMGs between mon1 and mon48 (KRT71, CD44, ROR2 and ZDHHC13) were further validated by bisulfite sequencing. Conclusions This study revealed dynamic changes in the genomic methylation profiles of mon1, mon24, and mon48 sheep, and the percentages of methylated cytosines were 3.38%, 2.85% and 4.17%, respectively. Of the DMGs, KRT71 and CD44 were highly methylated in mon1, and ROR2 and ZDHHC13 were highly methylated in mon48. These findings provide foundational information that may be used to develop strategies for potentially retaining the lamb fur and thus improving the economic value of Tan sheep. Supplementary Information The online version contains supplementary material available at 10.1186/s40104-021-00632-9.

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Integrated miRNA-mRNA analysis reveals regulatory pathways underlying the curly fleece trait in Chinese tan sheep.

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Integrated miRNA-mRNA analysis reveals regulatory pathways underlying the curly fleece trait in Chinese tan sheep.

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