Target Genes Of Differentially Expressed miRNAs Research Articles

Identification and comparative analysis of miRNA transcriptomes after allograft and xenograft transplantation in Pinctada fucata martensii

Effective immune regulation after transplantation during pearl production is crucial for the cultivation of high-quality pearls. MicroRNAs (miRNAs) play an important role in a variety of physiological processes. To understand the regulatory rules of miRNAs after transplantation in Pinctada funcata martensii, we constructed 13 miRNA transcriptomes, including the control group (Con), allograft (Al), and xenograft (Xe) transplantation at six time points (6, 12, and 24 h and 3, 6, and 12 days), in which the xenografted mantle tissue was from Pinctada maxima. We identified 159 differentially expressed miRNAs (DEMs) and found that these DEMs showed high expression at 12 h, 24 h, and 3 days after transplantation. A total of 130 DEMs, such as Let-7, were present in the Al and Xe groups; miR-34 and 16 other DEMs were specifically present in the Al group; miR-216b and 13 other DEMs were specifically present in the Xe group. Compared with the Con group, the target genes of DEMs in the Al group were significantly enriched in protein complex, cytoskeleton, and macromolecular complex, and the Xe group was significantly enriched in ribonucleoside metabolic process, nucleoside binding, and cell division. Compared with the Al group, the target genes in the Xe group were significantly enriched in response to DNA damage stimulation. Overall, multiple pathways associated with cellular activity were enriched in higher numbers of genes in the Xe group than in the Al group. These findings enriched the information on immune regulatory mechanisms at the expression level of miRNAs in P. f. martensii after transplantation.

A preliminary integrated analysis of miRNA-mRNA expression profiles reveals a role of miR-146a-3p/TRAF6 in plasma from gestational diabetes mellitus patients.

To utilize an integrative strategy to construct functional miRNA-mRNA regulatory networks by combining the reverse expression relationships between miRNAs and targets and computational predictions for gestational diabetes mellitus (GDM). A total of three microarray or RNA-seq datasets (GSE98043, GSE19649 and GSE92772) of plasma samples comparing GDM pregnant women and healthy control pregnant women were downloaded from the GEO database. The differentially expressed genes (DEmRNAs) and the differentially expressed miRNAs (DEmiRNAs) were analyzed. The target genes of DEmiRNAs were identified using two independent and complementary types of information: computational target predictions and expression relationships. An interaction network was constructed to identify hub genes of GDM. Another dataset (GSE92772) was used to externally verify the predictive ability of the hub genes. A total of 264 DEmiRNAs and 1217 DEmRNAs were identified with Hsa-miR-146a-3p ranked first of DEmiRNAs. Functions of GDM-related miRNAs were mainly enriched in the glypican pathway, proteoglycan syndecan-mediated signaling events, and syndecan-1-mediated signaling events. A total of 47 target genes, including TRAF6, were shared between the computational target predictions and DEmRNAs and were identified as target genes of hsa-miR-146a-3p. The interaction network analysis identified TRAF6, CASP8, PTPN6, and CHD3 as hub genes involved in the pathophysiological process of GDM. Next, independent external validation was performed using the GSE19649 dataset. The expression of TRAF6, CASP8 and CHD3 in eight pairs of GDM blood samples was confirmed to be higher than that in healthy pregnant women blood samples with a AUC of 0.813, 0.813, and 0.703, respectively. Our preliminary analysis revealed that miR-146a-3p/TRAF6 might play a central role in the pathogenesis of GDM. Three hub genes, TRAF6, CASP8, and CHD3, were identified and independently externally validated as potential GDM noninvasive serum markers for future biomarkers research.

Differential expression profile of miRNAs between stable and vulnerable plaques of carotid artery stenosis patients.

Plaque vulnerability is associated with the degree of carotid artery stenosis (CS) and the risk of stroke. MicroRNAs (miRNAs) exert critical functions in disease progression, although only a few miRNAs have been well identified in CS. Therefore, this study aimed to investigate the differential expression profile of miRNAs and their potential functions in plaques of CS patients. Three CS patients with stable plaques and three patients with vulnerable plaques who underwent carotid endarterectomy were enrolled in this study. Differentially expressed miRNAs (DEmiRNAs) between patients with stable and vulnerable plaques were determined using small RNA sequencing. Target genes of DEmiRNAs were predicted and submitted to functional analyses. Validation of dysregulated DEmiRNAs was determined using quantitative real-time polymerase chain reaction (qRT-PCR). After sequencing, 76 DEmiRNAs were identified in vulnerable plaques, including 53 upregulated miRNAs and 23 downregulated miRNAs. Next, 23,495 target genes of the identified DEmiRNAs were predicted and functionally analyzed. This indicated that the target genes of the identified DEmiRNAs were mainly enriched in protein phosphorylation, transcription, nitrogen compound metabolism, endocytosis and autophagy, and related to signaling pathways of Hippo, MAPK, insulin, TGF-β, FoxO, AMPK and p53. Furthermore, qRT-PCR results for six miRNAs showed that five (83%) of them (hsa-miR-511-5p, hsa-miR-150-5p, hsa-miR-378a-5p, hsa-miR-365b-5p and hsa-miR-6511b-5p) were consistent with the sequencing results. Differential expression profiles and potential function of miRNAs associated with plaque stability in CS patients are identified for the first time, which should help to understand the regulatory mechanism of plaque stability in CS.

Molecular characterization of extracellular vesicles derived from follicular fluid of women with and without PCOS: integrating analysis of differential miRNAs and proteins reveals vital molecules involving in PCOS

PurposeTo elucidate the characterization of extracellular vesicles (EVs) in the follicular fluid-derived extracellular vesicles (FF-EVs) and discover critical molecules and signaling pathways associating with the etiology and pathobiology of PCOS, the differentially expressed miRNAs (DEmiRNAs) and differentially expressed proteins profiles (DEPs) were initially explored and combinedly analyzed.MethodsFirst, the miRNA and protein expression profiles of FF-EVs in PCOS patients and control patients were compared by RNA-sequencing and tandem mass tagging (TMT) proteomic methods. Subsequently, Gene Ontology and the Kyoto Encyclopedia of Genes and Genomes were used to analyze the biological function of target genes of DEmiRNAs and DEPs. Finally, to discover the functional miRNA-target gene-protein interaction pairs involved in PCOS, DEmiRs target gene datasets and DEPs datasets were used integratedly.ResultsA total of 6 DEmiRNAs and 32 DEPs were identified in FF-EVs in patients with PCOS. Bioinformatics analysis revealed that DEmiRNAs target genes are mainly involved in thiamine metabolism, insulin secretion, GnRH, and Apelin signaling pathway, which are closely related to the occurrence of PCOS. DEPs also closely related to hormone metabolism processes such as steroid hormone biosynthesis. In the analysis integrating DEmiRNAs target genes and DEPs, two molecules, GRAMD1B and STPLC2, attracted our attention that are closely associated with cholesterol transport and ceramide biosynthesis, respectively.ConclusionDysregulated miRNAs and proteins in FF-EVs, mainly involving in hormone metabolism, insulin secretion, neurotransmitters regulation, adipokine expression, and secretion, may be closely related to PCOS. The effects of GRAMD1B and STPLC2 on PCOS deserve further study.

Identification and analysis of key microRNAs derived from osteoarthritis synovial fluid exosomes.

Identification and analysis of key microRNAs derived from osteoarthritis synovial fluid exosomes.

LncRNA HIF1A-AS1 Regulates the Cellular Function of HUVECs by Globally Regulating mRNA and miRNA Expression.

Long non-coding RNA (lncRNA) hypoxia inducible factor 1α-antisense RNA 1 (HIF1A-AS1) serves critical roles in cardiovascular diseases (CVDs). Vascular endothelial cells (VECs) are vulnerable to stimuli. Our previous study revealed that knockdown of HIF1A-AS1 reduces palmitic acid-induced apoptosis and promotes the proliferation of human VECs (HUVECs); however, the underlying mechanism remains unclear. Cell Counting Kit-8, flow cytometry, transwell invasion, and wound healing were applied to detect the function of HUVECs. Moreover, miRNA sequencing (miRNA-seq) and RNA sequencing (RNA-seq) were conducted to uncover its underlying mechanism. Quantitative Polymerase Chain Reaction (qPCR) was implemented to assess the accuracy of miRNA-seq. A co-expression network was generated to determine the relationship between differentially expressed miRNAs (DEmiRNAs) and differentially expressed genes (DEGs). Knockdown of HIF1A-AS1 promoted the proliferation, migration, and invasion but reduced the apoptosis of HUVECs, and the overexpression of this lncRNA had the opposite effect. Numerous DEmiRNAs and DEGs were identified, which might contribute to this phenomenon. Multiple target genes of DEmiRNAs were associated with cell proliferation and apoptosis, and overlapped with DEGs identified from RNA-seq. Finally, the network manifested that lncRNA HIF1A-AS1 moderated the function of HUVECs by not only regulating the expression of some genes directly but also by influencing a few miRNAs to indirectly mediate the expression of mRNAs. The results suggested that HIF1A-AS1 might regulate HUVEC function by not only regulating the expression of some genes directly but also by influencing some miRNAs to indirectly mediate the expression level of mRNA.

High-Throughput Sequencing Reveals Novel microRNAs Involved in the Continuous Flowering Trait of Longan (Dimocarpus longan Lour.).

A major determinant of fruit production in longan (Dimocarpus longan Lour.) is the difficulty of blossoming. In this study, high-throughput microRNA sequencing (miRNA-Seq) was carried out to compare differentially expressed miRNAs (DEmiRNAs) and their target genes between a continuous flowering cultivar 'Sijimi' (SJ), and a unique cultivar 'Lidongben' (LD), which blossoms only once in the season. Over the course of our study, 1662 known miRNAs and 235 novel miRNAs were identified and 13,334 genes were predicted to be the target of 1868 miRNAs. One conserved miRNA and 29 new novel miRNAs were identified as differently expressed; among them, 16 were upregulated and 14 were downregulated. Through the KEGG pathway and cluster analysis of DEmiRNA target genes, three critical regulatory pathways, plant-pathogen interaction, plant hormone signal transduction, and photosynthesis-antenna protein, were discovered to be strongly associated with the continuous flowering trait of the SJ. The integrated correlation analysis of DEmiRNAs and their target mRNAs revealed fourteen important flowering-related genes, including COP1-like, Casein kinase II, and TCP20. These fourteen flowering-related genes were targeted by five miRNAs, which were novel-miR137, novel-miR76, novel-miR101, novel-miR37, and csi-miR3954, suggesting these miRNAs might play vital regulatory roles in flower regulation in longan. Furthermore, novel-miR137 was cloned based on small RNA sequencing data analysis. The pSAK277-miR137 transgenic Arabidopsis plants showed delayed flowering phenotypes. This study provides new insight into molecular regulation mechanisms of longan flowering.

Integrated Transcriptome Analysis Reveals the Crucial mRNAs and miRNAs Related to Fecundity in the Hypothalamus of Yunshang Black Goats during the Luteal Phase.

A normal estrus cycle is essential for the breeding of goats, and the luteal phase accounts for most of the estrus cycle. The corpus luteum (CL) formed during the luteal phase is a transient endocrine gland that is crucial for the reproductive cycle and pregnancy maintenance, and is controlled by many regulatory factors. However, the molecular mechanism of the hypothalamus effect on the reproductive performance of different litter sizes during the luteal phase of goats has not been elucidated. In this study, RNA-sequencing was used to analyze the mRNA and miRNA expression profiles of the hypothalamic tissues with the high-fecundity goats during the luteal phase (LP-HF) and low-fecundity goats during the luteal phase (LP-LF). The RNA-seq results found that there were 1963 differentially expressed genes (DEGs) (890 up-regulated and 1073 down-regulated). The miRNA-seq identified 57 differentially expressed miRNAs (DEMs), including 11 up-regulated and 46 down-regulated, of which 199 DEGs were predicted to be potential target genes of DEMs. Meanwhile, the functional enrichment analysis identified several mRNA-miRNA pairs involved in the regulation of the hypothalamic activity, such as the common target gene MEA1 of novel-miR-972, novel-miR-125 and novel-miR-403, which can play a certain role as a related gene of the reproductive development in the hypothalamic-pituitary-gonadal (HPG) axis and its regulated network, by regulating the androgen secretion. While another target gene ADIPOR2 of the novel-miR-403, is distributed in the hypothalamus and affects the reproductive system through a central role on the HPG axis and a peripheral role in the gonadal tissue. An annotation analysis of the DE miRNA-mRNA pairs identified targets related to biological processes, such as anion binding (GO:0043168) and small molecule binding (GO: 0036094). Subsequently, the KEGG(Kyoto Encyclopedia of Genes and Genomes) pathways were performed to analyze the miRNA-mRNA pairs with negatively correlated miRNAs. We found that the GnRH signaling pathway (ko04912), the estrogen signaling pathway (ko04915), the Fc gamma R-mediated phagocytosis (ko04666), and the IL-17 signaling pathway (ko04657), etc., were directly and indirectly associated with the reproductive process. These targeting interactions may be closely related to the reproductive performance of goats. The results of this study provide a reference for further research on the molecular regulation mechanism for the high fertility in goats.

Analysis of circRNA and miRNA expression profiles in IGF3-induced ovarian maturation in spotted scat (Scatophagus argus).

Insulin-like growth factor 3 (IGF3) induces ovarian maturation in teleosts; however, research on its molecular regulatory mechanism remains deficient. Circular RNAs (circRNAs) and microRNAs (miRNAs) are involved in various biological processes, including reproduction. In this study, circRNAs and miRNAs involved in IGF3-induced ovarian maturation were evaluated in spotted scat (Scatophagus argus). In ovarian tissues, we identified 176 differentially expressed (DE) circRNAs and 52 DE miRNAs between IGF3 treatment and control groups. Gene Ontology (GO) enrichment analyses showed that host genes of DE circRNAs and target genes of DE miRNAs were enriched for various processes with a high degree of overlap, including cellular process, reproduction, reproductive process, biological adhesion, growth, extracellular region, cell junction, catalytic activity, and transcription factor activity. Enriched Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways included cell adhesion molecules, ECM-receptor interaction, regulation of actin cytoskeleton, focal adhesion, cell cycle, Hedgehog signaling pathway, phosphatidylinositol signaling system, PI3K-Akt signaling pathway, Apelin signaling pathway, Notch signaling pathway, insulin signaling pathway, and Rap1 signaling pathway. A circRNA-miRNA-mRNA regulatory network was constructed, including DE genes involved in reproduction (e.g., oocyte maturation, oocyte meiosis, and ECM remodeling), such as ccnd2, hecw2, dnm2, irs1, adam12, and cdh13. According to the regulatory network and tissue distribution, we identified one circRNA (Lachesis_group5:6245955|6270787) and three miRNAs (novel_miR_622, novel_miR_980, and novel_miR_64) that may exert regulatory effects in IGF3-induced ovarian maturation in S. argus. Taken together, this study provides a novel insight into the molecular mechanisms by which IGF3 functions in ovaries and highlights the effects of circRNAs and miRNAs in reproduction in S. argus.

Transcriptional analysis of microRNAs related to unsaturated fatty acid synthesis by interfering bovine adipocyte ACSL1 gene.

Long-chain fatty acyl-CoA synthase 1 (ACSL1) plays a vital role in the synthesis and metabolism of fatty acids. The proportion of highly unsaturated fatty acids in beef not only affects the flavor and improves the meat’s nutritional value. In this study, si-ACSL1 and NC-ACSL1 were transfected in bovine preadipocytes, respectively, collected cells were isolated on the fourth day of induction, and then RNA-Seq technology was used to screen miRNAs related to unsaturated fatty acid synthesis. A total of 1,075 miRNAs were characterized as differentially expressed miRNAs (DE-miRNAs), of which the expressions of 16 miRNAs were upregulated, and that of 12 were downregulated. Gene ontology analysis indicated that the target genes of DE-miRNAs were mainly involved in biological regulation and metabolic processes. Additionally, KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis identified that the target genes of DE-miRNAs were mainly enriched in metabolic pathways, fatty acid metabolism, PI3K-Akt signaling pathway, glycerophospholipid metabolism, fatty acid elongation, and glucagon signaling pathway. Combined with the previous mRNA sequencing results, several key miRNA-mRNA targeting relationship pairs, i.e., novel-m0035-5p—ACSL1, novel-m0035-5p—ELOVL4, miR-9-X—ACSL1, bta-miR-677—ACSL1, miR-129-X—ELOVL4, and bta-miR-485—FADS2 were screened via the miRNA-mRNA interaction network. Thus, the results of this study provide a theoretical basis for further research on miRNA regulation of unsaturated fatty acid synthesis in bovine adipocytes.

Small RNA sequencing reveals sex-related miRNAs in Collichthys lucidus.

Collichthys lucidus (C. lucidus) is an economically important fish species, exhibiting sexual dimorphism in its growth rate. However, there is a lack of research on its underlying sex-related mechanisms. Therefore, small RNA sequencing was performed to better comprehend these sex-related molecular mechanisms. In total, 171 differentially expressed miRNAs (DE-miRNAs) were identified between the ovaries and testes. Functional enrichment analysis revealed that the target genes of DE-miRNAs were considerably enriched in the p53 signaling, PI3K–Akt signaling, and TGF-beta signaling pathways. In addition, sex-related miRNAs were identified, and the expression of miR-430c-3p and miR-430f-3p was specifically observed in the gonads compared with other organs and their expression was markedly upregulated in the testes relative to the ovaries. Bmp15 was a target of miR-430c-3p and was greatly expressed in the ovaries compared with the testes. Importantly, miR-430c-3p and bmp15 co-expressed in the ovaries and testes. This research provides the first detailed miRNA profiles for C. lucidus concerning sex, likely laying the basis for further studies on sex differentiation in C. lucidus.

Integrated microarray for identifying the hub mRNAs and constructed miRNA-mRNA network in coronary in-stent restenosis.

As a major complication after percutaneous coronary intervention (PCI) in patients who suffer from coronary artery disease, in-stent restenosis (ISR) poses a significant challenge for clinical management. A miRNA-mRNA regulatory network of ISR can be constructed to better reveal the occurrence of ISR. The relevant data set from the Gene Expression Omnibus (GEO) database was downloaded, and 284 differentially expressed miRNAs (DE-miRNAs) and 849 differentially expressed mRNAs (DE-mRNAs) were identified. As predicted by online tools, 65 final functional genes (FmRNAs) were overlapping DE-mRNAs and DE-miRNAs target genes. In the biological process (BP) terms of gene ontology (GO) functional analysis, the FmRNAs were mainly enriched in the cellular response to peptide, epithelial cell proliferation, and response to peptide hormone. In the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, the FmRNAs were mainly enriched in breast cancer, endocrine resistance, and Cushing syndrome. Jun proto-oncogene, activator protein-1 (AP-1) transcription factor subunit (JUN), insulin-like growth factor 1 receptor (IGF1R), member RAS oncogene family (RAB14), specificity protein 1 (SP1), protein tyrosine phosphatase nonreceptor type 1 (PTPN1), DDB1 and CUL4 associated factor 10 (DCAF10), retinoblastoma-binding protein 5 (RBBP5), and eukaryotic initiation factor 4A-I (EIF4A1) were hub genes in the protein-protein interaction network (PPI network). The miRNA-mRNA network containing DE-miRNAs and hub genes was built. Hsa-miR-139-5p-JUN, hsa-miR-324-5p-SP1 axis pairs were found in the miRNA-mRNA network, which could promote ISR development. The aforementioned results indicate that the miRNA-mRNA network constructed in ISR has a regulatory role in the development of ISR and may provide new approaches for clinical treatment and experimental development.

Pathogenesis and Therapeutic Targets of Focal Cortical Dysplasia Based on Bioinformatics Analysis.

Focal cortical dysplasia (FCD), a malformation of cortical development, is the most common cause of intractable epilepsy in children. However, the causes and underlying molecular events of FCD need further investigation. The microarray dataset GSE62019 and GSE97365 were obtained from Gene Expression Omnibus. To examine critical genes and signaling pathways, bioinformatics analysis tools such as protein-protein interaction (PPI) networks, miRNA-mRNA interaction networks, and immune infiltration in FCD samples were used to fully elucidate the pathogenesis of FCD. A total of 534 differentially expressed genes (DEGs) and 71 differentially expressed miRNAs (DEMs) were obtained. The DEGs obtained were enriched in ribosomal, protein targeting, and pathways of neurodegeneration multiple diseases, whereas the target genes of DEMs were enriched in signaling pathways such as transforming growth factor beta, Wnt, PI3K-Akt, etc. Finally, four hub genes (RPL11, FAU, RPS20, RPL27) and five key miRNAs (hsa-let-7b, hsa-miR-185, hsa-miR-23b, hsa-miR-222 and hsa-miR-92b) were obtained by PPI network, miRNA-mRNA network, and ROC analysis. The immune infiltration results showed that the infiltration levels of five immune cells (MDSC, regulatory T cells, activated CD8+ T cells, macrophage and effector memory CD8+ T cells) were slightly higher in FCD samples than in control samples. Moreover, the gene expressions of RPS19, RPL19, and RPS24 were highly correlated with the infiltration levels and immune characteristics of 28 immune cells. It broadens the understanding of the molecular mechanisms underlying the development of FCD and enlightens the identification of molecular targets and diagnostic biomarkers for FCD.

Prmt7 Downregulation in Mouse Spermatogonia Functions through miR-877-3p/Col6a3.

Protein arginine methyltransferases 7 (Prmt7) is expressed in male germ cells, including primordial germ cells, gonocytes, and spermatogonia. Our previous study demonstrated that Prmt7 downregulation reduced the proliferation of GC-1 cells (a cell line of mouse immortalized spermatogonia). However, how Prmt7 regulates spermatogonial proliferation through miRNA and the target gene remains elusive. Here, we experimentally reduced the Prmt7 expression in the GC-1 cells and subjected them to miRNA sequencing to explore the miRNA profile and its Prmt7-responsive members. In total, 48 differentially expressed miRNAs (DEmiRNAs), including 36 upregulated and 12 downregulated miRNAs, were identified. After verifying the validity of sequencing results through qRT-PCR assays in randomly selected DEmiRNAs, we predicted the target genes of these DEmiRNAs. Next, we combined DEmiRNA target genes and previously identified differentially expressed genes between Prmt7 knockdown and control groups of GC-1 cells, which resulted in seven miRNA/target gene pairs. Among these miRNA/target gene pairs, we further detected the expression of Col6a3 (collagen type VI alpha 3) as the target gene of mmu-miR-877-3p. The results suggested that Prmt7 downregulation in mouse spermatogonia might function through miR-877-3p/Col6a3. Overall, these findings provide new insights into the role of Prmt7 in male germ cell development through miRNA and target genes.

Exosome-transmitted miR-3124-5p promotes cholangiocarcinoma development via targeting GDF11.

ObjectiveCholangiocarcinoma (CHOL) is a deadly cancer worldwide with limited available therapies. The aim of this study was to investigate key exosomal miRNAs and their functions in CHOL development.MethodsSerum exosomes were isolated from patients with CHOL and healthy controls, followed by miRNA sequencing for identifying differentially expressed miRNAs (DEMs) and their functions. Then, the expression of key DEMs was experimentally validated in exosomes from clinical CHOL patients and CHOL cells. The effects of overexpression of key DEMs on CHOL cell migration and proliferation were investigated. A key exosomal DEM miR-3124-5p was identified. The effects of overexpression or knockdown of exosomal miR-3124-5p on the proliferation, migration, and angiogenesis of human umbilical vein endothelial cells (HUVECs) were investigated. Moreover, the function of exosomal miR-3124-5p on tumor growth in vivo was explored.ResultsA total of 632 exosomal DEMs were identified between CHOL and control samples. Target genes of DEMs were significantly enriched in pathways, such as the p53 signaling pathway. miR-3124-5p was upregulated in serum exosomes from CHOL patients and exosomes from CHOL cells, and overexpression of miR-3124-5p promoted RBE cell migration and viability. Moreover, overexpression of exosomal miR-3124-5p promoted the proliferation, migration, and angiogenesis of HUVECs, while knockdown of miR-3124-5p had the opposite effect. miR-3124-5p could target growth differentiation factor 11 (GDF11) and downregulate GDF11 expression. Furthermore, exosomal miR-3124-5p promoted tumor growth in vivo.ConclusionsOur findings revealed that exosome-encapsulated miR-3124-5p promoted the malignant progression of CHOL by targeting GDF11. Exosomal miR-3124-5p and GDF11 could be promising biomarkers or therapeutic targets for CHOL.

Transcriptome-wide study revealed m6A and miRNA regulation of embryonic breast muscle development in Wenchang chickens.

N6-Methyladenosine (m6A) modification has been shown to play important role in skeletal muscle development. Wenchang chickens are commonly used as a high-quality animal model in researching meat quality. However, there have been no previous reports regarding the profile of m6A and its function in the embryonic breast muscle development of Wenchang chickens. In this paper, we identified different developmental stages of breast muscle in Wenchang chickens and performed m6A sequencing and miRNA sequencing in the breast muscle of embryos. Embryo breast muscles were weighed and stained with hematoxylin–eosin after hatching. We found that myofibers grew fast on the 10th day after hatching (E10) and seldom proliferated beyond the 19th day after hatching (E19). A total of 6,774 differentially methylated genes (DMGs) were identified between E10 and E19. For RNA-seq data, we found 5,586 differentially expressed genes (DEGs). After overlapping DEGs and DMGs, we recorded 651 shared genes (DEMGs). Subsequently, we performed miRNA-seq analysis and obtained 495 differentially expressed miRNAs (DEMs). Then, we overlapped DEMGs and the target genes of DEMs and obtained 72 overlapped genes (called miRNA-m6A-genes in this study). GO and KEGG results showed DEMGs enriched in many muscle development-related pathways. Furthermore, we chose WNT7B, a key regulator of skeletal muscle development, to perform IGV visualization analysis and found that the m6A levels on the WNT7B gene between E10 and E19 were significantly different. In conclusion, we found that miRNAs, in conjunction with m6A modification, played a key role in the embryonic breast muscle development of Wenchang chickens. The results of this paper offer a theoretical basis for the study of m6A function in muscle development and fat deposition of Wenchang chickens.

Differential MicroRNA Expression in Porcine Endometrium Related to Spontaneous Embryo Loss during Early Pregnancy.

Litter size is an important indicator to measure the production capacity of commercial pigs. Spontaneous embryo loss is an essential factor in determining sow litter size. In early pregnancy, spontaneous embryo loss in porcine is as high as 20–30% during embryo implantation. However, the specific molecular mechanism underlying spontaneous embryo loss at the end of embryo implantation remains unknown. Therefore, we comprehensively used small RNA sequencing technology, bioinformatics analysis, and molecular experiments to determine the microRNA (miRNA) expression profile in the healthy and arresting embryo implantation site of porcine endometrium on day of gestation (DG) 28. A total of 464 miRNAs were identified in arresting endometrium (AE) and healthy endometrium (HE), and 139 differentially expressed miRNAs (DEMs) were screened. We combined the mRNA sequencing dataset from the SRA database to predict the target genes of these miRNAs. A quantitative real-time PCR assay identified the expression levels of miRNAs and mRNAs. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were performed on differentially expressed target genes of DEMs, mainly enriched in epithelial development and amino acids metabolism-related pathways. We performed fluorescence in situ hybridization (FISH) and the dual-luciferase report gene assay to confirm miRNA and predicted target gene binding. miR-205 may inhibit its expression by combining 3′-untranslated regions (3′ UTR) of tubulointerstitial nephritis antigen-like 1 (TINAGL1). The resulting inhibition of angiogenesis in the maternal endometrium ultimately leads to the formation of arresting embryos during the implantation period. This study provides a reference for the effect of miRNA on the successful implantation of pig embryos in early gestation.

Differential expression of miRNAs in the body wall of the sea cucumber Apostichopus japonicus under heat stress.

MicroRNAs, as one of the post-transcriptional regulation of genes, play an important role in the development process, cell differentiation and immune defense. The sea cucumber Apostichopus japonicus is an important cold-water species, known for its excellent nutritional and economic value, which usually encounters heat stress that affects its growth and leads to significant economic losses. However, there are few studies about the effect of miRNAs on heat stress in sea cucumbers. In this study, high-throughput sequencing was used to analyze miRNA expression in the body wall of sea cucumber between the control group (CS) and the heat stress group (HS). A total of 403 known miRNAs and 75 novel miRNAs were identified, of which 13 miRNAs were identified as significantly differentially expressed miRNAs (DEMs) in response to heat stress. A total of 16,563 target genes of DEMs were predicted, and 101 inversely correlated target genes that were potentially regulated by miRNAs in response to heat stress of sea cucumbers were obtained. Based on these results, miRNA-mRNA regulatory networks were constructed. The expression results of high-throughput sequencing were validated in nine DEMs and four differentially expressed genes (DEGs) by quantitative real-time polymerase chain reaction (RT-qPCR). Moreover, pathway enrichment of target genes suggested that several important regulatory pathways may play an important role in the heat stress process of sea cucumber, including ubiquitin-mediated proteolysis, notch single pathway and endocytosis. These results will provide basic data for future studies in miRNA regulation and molecular adaptive mechanisms of sea cucumbers under heat stress.

Construction of a miRNA-mRNA Network Related to Exosomes in Colon Cancer

Background The competing endogenous RNA (CeRNA) network plays important roles in the occurrence and development of colon cancer. This research is aimed at constructing a miRNA-mRNA network associated with exosomes in colon cancer. Methods We explored the GEO database and then analyzed the RNAs of 722 samples to obtain differentially expressed miRNAs (DEMs) and mRNAs (DEGs) alongside the progress of colon cancer. Next, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of DEM target genes and DEGs were performed. In addition, a miRNA-mRNA network related to exosomes in colon cancer was constructed based on DEMs and DEGs. Finally, the expression of miRNA and mRNA in the network was verified by GEPIA2 on the base of TCGA database. Results Through our analysis, 19 DEMs (17 up and 2 down) and 1672 DEGs (954 up and 718 down) were screened. The GO and KEGG results show that these DEGs were mainly enriched in ribonucleoprotein complex biogenesis, noncoding RNA metabolic process, cell-substrate junction, cadherin binding, transcription coregulator activity, and regulation of the human T-cell leukemia virus 1 infection-related pathway. Besides, a miRNA-mRNA network, including 4 miRNAs (hsa-miR-623, hsa-miR-320c, hsa-miR-486-5p, and hsa-miR-1290) and 7 mRNAs (GNAI1, CADM1, PGRMC2, etc.), was constructed. Three of these seven mRNAs were downregulated in colon cancer. Ultimately, the GNAI1, CADM1, and PGRMC2 expression levels were verified by TCGA database. Conclusions This study reveals the network relationship between colon cancer exosome-derived miRNA and targeted mRNA. It deepens our understanding of new molecular mechanisms and pathways that may play a role in the occurrence and metastasis of colon cancer.

Genome-wide identification of chicken bursae of Fabricius miRNAs in response to very virulent infectious bursal disease virus.

Infectious bursal disease virus (IBDV) can cause a highly contagious immunosuppressive disease in young chickens. MicroRNAs (miRNAs) are crucial regulators of gene expression and are involved in the pathogenesis of IBDV infection. To investigate the roles of miRNA in chicken bursae of Fabricius in response to very virulent IBDV (vvIBDV) infection, RNA sequencing was performed to compare the small RNA libraries from uninfected and vvIBDV-infected group which was infected for 3 days. A total of 77 differentially expressed (DE) miRNAs were identified in BF, of which 42 DE miRNAs were upregulated and 35 DE miRNAs were downregulated. A gene ontology analysis showed that genes associated with cellular processes, cells, and binding were enriched. Moreover, pathway analyses suggested that apoptosis, T cell receptor signaling pathways, and chemokine signaling pathways may be activated following vvIBDV infection. In addition, we predicted the target genes of DE miRNAs and constructed an miRNA-mRNA regulatory network. In total, 189 pairs of miRNA-target genes were identified, comprising 67 DE miRNAs and 73 mRNAs. In this network, gga-miR-1684b-3p was identified with the highest fold change, as well as gga-miR-1788-3p and gga-miR-3530-5p showed a high degree of change. The above three miRNAs were considered to play vital roles in vvIBDV-host interactions. This study was the first to perform a comprehensive analysis of DE miRNAs in the bursa of Fabricius in response to vvIBDV infection, and it provided new insights into molecular mechanisms underlying vvIBDV infection and pathogenesis.