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      Identification and characterization of microRNAs in the intestinal tissues of sheep ( Ovis aries)

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          Abstract

          Sheep are small ruminants, and their long intestines exhibit high digestive and absorptive capacity in many different rearing conditions; however, the genetic bases of this characteristic remains unclear. MicroRNAs (miRNAs) play a major role in maintaining both intestinal morphological structure as well as in regulating the physiological functions of this organ. However, no study has reported on the miRNA expression profile in the intestinal tissues of sheep. Here, we analyzed and identified the miRNA expression profile of three different intestinal tissues (i.e., duodenum, cecum, and colon) of sheep ( Ovis aries) using high-throughput sequencing and bioinformatic methods. In total, 106 known miRNAs were identified, 458 conserved miRNAs were detected, 192 unannotated novel miRNAs were predicted, and 195 differentially expressed miRNAs were found between the different tissues. Additionally, 3,437 candidate target genes were predicted, and 17 non-redundant significantly enriched GO terms were identified using enrichment analysis. A total of 99 candidate target genes were found to significantly enriched in 4 KEGG biological pathways. A combined regulatory network was constructed based on 92 metabolism-related candidate target genes and 65 differentially expressed miRNAs, among which 7 miRNAs were identified as hub miRNAs. Via these mechanisms, miRNAs may play a role in maintaining intestinal homeostasis and metabolism. This study helps to further explain the mechanisms that underlie differences in tissue morphology and function in three intestinal segments of sheep.

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          Prediction of mammalian microRNA targets.

          MicroRNAs (miRNAs) can play important gene regulatory roles in nematodes, insects, and plants by basepairing to mRNAs to specify posttranscriptional repression of these messages. However, the mRNAs regulated by vertebrate miRNAs are all unknown. Here we predict more than 400 regulatory target genes for the conserved vertebrate miRNAs by identifying mRNAs with conserved pairing to the 5' region of the miRNA and evaluating the number and quality of these complementary sites. Rigorous tests using shuffled miRNA controls supported a majority of these predictions, with the fraction of false positives estimated at 31% for targets identified in human, mouse, and rat and 22% for targets identified in pufferfish as well as mammals. Eleven predicted targets (out of 15 tested) were supported experimentally using a HeLa cell reporter system. The predicted regulatory targets of mammalian miRNAs were enriched for genes involved in transcriptional regulation but also encompassed an unexpectedly broad range of other functions.
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            Plasma extracellular RNA profiles in healthy and cancer patients

            Extracellular vesicles are selectively enriched in RNA that has potential as disease biomarkers. To systemically characterize circulating extracellular RNA (exRNA) profiles, we performed RNA sequencing analysis on plasma extracellular vesicles derived from 50 healthy individuals and 142 cancer patients. Of ~12.6 million raw reads for each individual, the number of mappable reads aligned to RNA references was ~5.4 million including miRNAs (~40.4%), piwiRNAs (~40.0%), pseudo-genes (~3.7%), lncRNAs (~2.4%), tRNAs (~2.1%), and mRNAs (~2.1%). By expression stability testing, we identified a set of miRNAs showing relatively consistent expression, which may serve as reference control for exRNA quantification. By performing multivariate analysis of covariance, we identified significant associations of these exRNAs with age, sex and different types of cancers. In particular, down-regulation of miR-125a-5p and miR-1343-3p showed an association with all cancer types tested (false discovery rate <0.05). We developed multivariate statistical models to predict cancer status with an area under the curve from 0.68 to 0.92 depending cancer type and staging. This is the largest RNA-seq study to date for profiling exRNA species, which has not only provided a baseline reference profile for circulating exRNA, but also revealed a set of RNA candidates for reference controls and disease biomarkers.
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              MicroRNAs in metabolism

              Abstract MicroRNAs (miRNAs) have within the past decade emerged as key regulators of metabolic homoeostasis. Major tissues in intermediary metabolism important during development of the metabolic syndrome, such as β‐cells, liver, skeletal and heart muscle as well as adipose tissue, have all been shown to be affected by miRNAs. In the pancreatic β‐cell, a number of miRNAs are important in maintaining the balance between differentiation and proliferation (miR‐200 and miR‐29 families) and insulin exocytosis in the differentiated state is controlled by miR‐7, miR‐375 and miR‐335. MiR‐33a and MiR‐33b play crucial roles in cholesterol and lipid metabolism, whereas miR‐103 and miR‐107 regulates hepatic insulin sensitivity. In muscle tissue, a defined number of miRNAs (miR‐1, miR‐133, miR‐206) control myofibre type switch and induce myogenic differentiation programmes. Similarly, in adipose tissue, a defined number of miRNAs control white to brown adipocyte conversion or differentiation (miR‐365, miR‐133, miR‐455). The discovery of circulating miRNAs in exosomes emphasizes their importance as both endocrine signalling molecules and potentially disease markers. Their dysregulation in metabolic diseases, such as obesity, type 2 diabetes and atherosclerosis stresses their potential as therapeutic targets. This review emphasizes current ideas and controversies within miRNA research in metabolism.
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                Author and article information

                Contributors
                Role: Project administrationRole: ResourcesRole: ValidationRole: Writing – original draftRole: Writing – review & editing
                Role: Writing – original draftRole: Writing – review & editing
                Role: Software
                Role: Validation
                Role: SoftwareRole: ValidationRole: Writing – original draftRole: Writing – review & editing
                Role: Formal analysisRole: Funding acquisitionRole: Project administrationRole: ResourcesRole: SoftwareRole: ValidationRole: Writing – original draftRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                28 February 2018
                2018
                : 13
                : 2
                : e0193371
                Affiliations
                [001]Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Shandong Agricultural University, Taian City, Shandong Province, China
                Laboratoire de Biologie du Développement de Villefranche-sur-Mer, FRANCE
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Author information
                http://orcid.org/0000-0001-5678-8202
                Article
                PONE-D-17-12242
                10.1371/journal.pone.0193371
                5831392
                29489866
                27e74d92-4c00-4103-a8b3-12801cf11190
                © 2018 Hou et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 29 March 2017
                : 9 February 2018
                Page count
                Figures: 5, Tables: 1, Pages: 17
                Funding
                Funded by: Shandong Provincial Modern Agriculture Industry Technology System
                Award ID: SDAIT-0901101
                Award Recipient :
                Funded by: National Key Technology Support Program
                Award ID: 2015BAD03B05
                Award Recipient :
                Funded by: Study on Super-high Reproductive New Breed and Supporting technological system of Small-Tailed Han Sheep
                Award ID: SDLZ2015131x
                Award Recipient :
                Funded by: Funds of Shandong "Double Tops" Program
                Award ID: No. SYL2017YSTD12
                Award Recipient :
                This work was financially supported by the Funds of Shandong “Double Tops” Program (No. SYL2017YSTD12), Shandong Provincial Modern Agriculture Industry Technology System (no. SDAIT-0901101), National Key Technology Support Program (2015BAD03B05) and Study on Super-high Reproductive New Breed and Supporting technological system of Small-Tailed Han Sheep (no. SDLZ2015131x). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Biology and life sciences
                Genetics
                Gene expression
                Gene regulation
                MicroRNAs
                Biology and life sciences
                Biochemistry
                Nucleic acids
                RNA
                Non-coding RNA
                MicroRNAs
                Biology and Life Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Medicine and Health Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Biology and Life Sciences
                Organisms
                Eukaryota
                Animals
                Vertebrates
                Amniotes
                Mammals
                Ruminants
                Sheep
                Biology and Life Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Cecum
                Medicine and Health Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Cecum
                Biology and Life Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Colon
                Medicine and Health Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Colon
                Biology and Life Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Duodenum
                Medicine and Health Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Duodenum
                Biology and Life Sciences
                Genetics
                Gene Expression
                Gene Regulation
                Biology and Life Sciences
                Computational Biology
                Gene Regulatory Networks
                Biology and Life Sciences
                Genetics
                Gene Regulatory Networks
                Custom metadata
                All relevant data are within the paper and its Supporting Information files.

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