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MicroRNAs (miRNA) plays an important role in several mammalian biological regulatory processes by post-transcriptionally regulating gene expression. However, there is little information on the miRNAs involved in the photoperiodism pathway that controls seasonal activity. To enhance our knowledge on the effect of different photoperiod conditions on miRNA, we divided Kazakh sheep into two groups: one exposed to a long photoperiod (LP, 16L:8D) and another with exposed to a short photoperiod (SP, 8L:16D) under supplemental feeding conditions. Further we compared the related miRNAs and target genes between the two groups. Fifteen differentially expressed miRNAs were identified, which were associated with 310 regulatory pathways covering photoperiodism, reproductive hormones, and nutrition. The miR-136-GNAQ pair was selected and validated as a differentially expressed, and a dual-luciferase reporter assay showed that the negative feedback loop existed between them. Examination of the expression profile revealed that the GNAQ expression was low in the estrous females both under LP and SP conditions, but high expression of GNAQ was observed in the anestrous females under LP conditions. Moreover, functional analysis revealed that KISS1 and GnRH expression was upregulated when GNAQ expression was downregulated in the hypothalamic cells, whereas DIO2 and TSHB expression was downregulated. Thus, miR-136-GNAQ might act as a switch in the regulation of seasonal estrus under different photoperiod conditions. These findings further enrich our understanding of the relationship between miRNAs and seasonal regulation of reproductive activity. Furthermore, our study provides novel insights into the miRNA-mediated regulatory mechanisms for overcoming photoinhibition in the seasonally breeding mammals, such as Kazakh sheep. [ABSTRACT FROM AUTHOR]

To examine the possible miRNA molecular regulatory mechanisms during maternal uterine involution after delivery, we selected ovary and uterus tissues that are structurally connected as experimental materials. We employed Illumina HiSeq sequencing to screen and analyze the quantity and characteristics of miRNA in postpartum ewes in the methylergometrine-treated group and physiological saline control group. Results showed that 16 miRNAs were identified in the ovary libraries, including 4 known miRNAs and 12 novel miRNAs. In the uterus libraries, 54 miRNAs were identified, which included 5 known miRNAs and 49 novel miRNAs. At the same time, target gene prediction, GO annotation, and KEGG signaling pathway enrichment analysis were employed. We found that maternal uterine involution after delivery may involve two miRNA-target gene pairs, i.e., miRNA-200a- ZEB1 and YAP1. The YAP1/Hippo signaling pathway is used to construct an ovary–uterine axial regulatory mechanism to regulate the restoration of postpartum maternal uterine morphology and function. In view of this, the identification of miRNAs with significant differences in this study fills a gap in research on miRNAs associated with regulation of postpartum uterine recovery in ewes and provided an important reference for comprehensive understanding and in-depth research on the regulatory molecular network mechanism for postpartum uterine involution in small ruminants. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are widely expressed in different mammalian tissues and exert their biological effects through corresponding target genes. miRNA target genes can be rapidly and efficiently identified and screened by combining bioinformatics prediction and experimental validation. To investigate the possible molecular regulatory mechanisms involving miRNAs during uterine involution in postpartum ewes, we used Illumina HiSeq sequencing technology to screen for the number and characteristics of miRNAs in faster uterine involution and normal uterine involution group. A total of 118 differentially expressed miRNAs, including 33 known miRNAs and 85 new miRNAs, were identified in the hypothalamic library, whereas 54 miRNAs, including 5 known miRNAs and 49 new miRNAs, were identified in the uterine library. Screening with four types of gene prediction software revealed 73 target genes associated with uterine involution, and subsequently, GO annotation and KEGG pathway analysis were performed. The results showed that, in the hypothalamic–uterine axis, uterine involution in postpartum ewes might primarily involve two miRNA-target gene pairs, namely, miRNA-200a–PTEN and miRNA-133–FGFR1, which can participate in GnRH signal transduction in the upstream hypothalamus and in the remodeling process at the downstream uterus, through the PI3K–AKT signaling pathway to influence the recovery of the morphology and functions of the uterus during the postpartum period in sheep. Therefore, identification of differentially expressed miRNAs in this study fills a gap in the research related to miRNAs in uterine involution in postpartum ewes and provides an important reference point for a comprehensive understanding of the molecular mechanisms underlying the regulation of postpartum uterine involution in female livestock. [ABSTRACT FROM AUTHOR]

Seasonal estrus is a critical limiting factor for animal fecundity. However, estrus occurs in some seasonally estrous sheep in the non-breeding season, and this phenomenon involves changes in ovarian biology. Previous studies indicated that small RNAs, such as microRNAs (miRNAs), play important regulatory roles in ovarian biology. Differentially expressed miRNAs in the ovaries of estrous sheep were identified using Solexa sequencing technology. A total of 423 known miRNAs were identified in ovaries of estrous sheep in the breeding season and non-breeding season. In the comparison of these two groups, 48 miRNAs were identified that were differentially expressed between the two groups (including 5 up-regulated and 43 down-regulated miRNAs). KEGG pathway analysis revealed that the target genes of some differentially expressed miRNAs were involved in pathways related to reproductive hormone signaling and follicular development. Furthermore, the levels of estradiol (E2), progesterone (P4), luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were lower in anestrus sheep than in sheep during the breeding season. Upon combining the pathway enrichment analysis, target gene expression and hormone measurement results, we suggest that these differentially expressed miRNAs might influence ovarian activity in the non-breeding season by regulating the above pathways. The identification of miRNAs that are differentially expressed between ovines in the breeding season and non-breeding season will contribute to our understanding of the role of miRNAs in estrus regulation, and these data may provide a basis for regulating estrus in sheep during the non-breeding season. [ABSTRACT FROM AUTHOR]

Given the important role of nutritional status for reproductive performance, we aimed to explore the potential microRNA (miRNA)-mRNA pairs and their regulatory roles associated with nutritional status in seasonal reproducing sheep. Individual ewes were treated with and without 0.3 kg/day concentrates, and the body condition score, estrus rate, and related miRNAs and target genes were compared. A total of 261 differentially expressed miRNAs were identified, including 148 hypothalamus-expressed miRNAs and 113 ovary-expressed miRNAs, and 349 target genes were predicted to be associated with nutritional status and seasonal reproduction in sheep. Ultimately, the miR-200b- GNAQ pair was screened and validated as differentially expressed, and a dual luciferase reporter assay showed that miR-200b could bind to the 3′-untranslated region of GNAQ to mediate the hypothalamic-pituitary-ovarian axis. Thus, miR-200b and its target gene GNAQ likely represent an important negative feedback loop, providing a link between nutritional status and seasonal reproduction in sheep toward enhancing reproductive performance and productivity. [ABSTRACT FROM AUTHOR]

Estrous regulation in sheep has an important role in the sheep industry in improving production of meat and wool. It has been reported that an enhanced nutritional status can induce estrus resulting in an end of the anestrous season earlier than occurs in ewes with a lesser nutritional status. However, the endocrine and physiological mechanisms that induce the increased incidence of estrus remains unclear. In the present study, the differences in amounts and characteristics of miRNAs in ewes at estrus or during the anestrous season were screened by using the Illumina HiSeq sequencing technology. In total, 294 miRNAs, including 174 novel miRNA candidates, were identified in ewes with an enhanced nutritional status (OEN) through assessment of the OEN library for this group and 307 miRNAs including 186 novel miRNA candidates were identified in the ewes with a lesser nutritional status (OAN) through assessing the OAN library, among which there were nine conserved and 104 novel miRNAs in differential amounts between the two libraries. Based on poly (A) q-PCR, six miRNAs were assessed to verify the accuracy of the library database. Furthermore, the family of the known miRNAs, the target genes and related pathways were also analyzed. The results indicated that the nutritional status had important roles in estrous regulation in sheep. The PLA2G4D can directly regulate ovarian follicle development, or indirectly influence leptin secretion involved in the regulation of the reproductive endocrine and physiological systems during the anestrous season. The identification of significantly different miRNAs expanded the repertoire of sheep miRNAs that have been examined and could contribute to further studies on the molecular mechanism of regulation of initiation of estrous cycles in previously anestrous ewes as influenced by different nutritional status. [ABSTRACT FROM AUTHOR]

Mannose-binding lectin (MBL) glycoproteins in blood can selectively recognise lectins on the surface of bacteria, and play an important role in natural immunity. Micro RNAs (miRNAs) are key molecules that regulate gene expression at the post-transcriptional level in vivo , and their pathways are specific and effective. Previous studies indicate that small RNAs such as miRNAs perform regulatory roles in immunology. Herein, we investigated differential expression of miRNAs during MBL protein immunotherapy in sheep following treatment with different MBL genotypes (resistant and susceptible), and identified miRNAs linked to different target genes and pathways. RNA was extracted from liver tissue of resistant and susceptible sheep, miRNAs were identified by high-throughput sequencing, and differentially expressed miRNAs were analysed by SOAP to predict target genes and biological pathways. Results: Some miRNAs (oar-mir-143, oar-mir-10b, oar-mir-382, oar-mir-432 and oar-mir-379) were up-regulated, while others were down-regulated. GPATCH3 and DNAJC5 were predicted target genes of oar-mir-379, DMRT1 and GATA4 were linked to oar-mir-382, and oar-mir-432 was associated with STAT2, DMRT1 and ATG16L1. Identification of miRNAs differentially expressed in resistant and susceptible sheep may expand our understanding of miRNAs in immune regulation, and the role of MBL in innate immunity. [ABSTRACT FROM AUTHOR]