Functional prediction of miR-3144-5p in human cardiac myocytes based on transcriptome sequencing and bioinformatics.
ABSTRACT: RAN guanine nucleotide release factor (RANGRF) encoding protein MOG1 plays an important role in cardiac arrhythmia, so we intended to investigate the regulatory miRNA of RANGRF and explore its potential regulatory mechanism in arrhythmogenesis.Based on bioinformatic analysis, miR-3144-5p was predicted to be a regulatory miRNA of RANGRF, which were then validated through a dual-luciferase reporter plasmid assay. Subsequently, the expression level of miR-3144-5p in human cardiac myocytes (HCMs) was detected, followed by cell transfection with miR-3144-5p mimics. Transcriptome sequencing was then performed in HCMs with or without transfection. The sequencing results were subjected to bioinformatic analyses, including differentially expressed gene (DEG) analysis, functional enrichment analysis, protein-protein interaction (PPI) network analysis, miRNA-target gene analysis, and miRNA-transcription factor (TF)-target gene coregulatory network analysis.There really existed a regulatory relation between miR-3144-5p and RANGRF. The expression level of miR-3144-5p was low in HCMs. After cell transfection, miR-3144-5p expression level significantly increased in HCMs. Bioinformatic analyses of the transcriptome sequencing results identified 300 upregulated and 271 downregulated DEGs between miR-3144-5p mimic and control group. The upregulated genes ISL1 and neuregulin 1 (NRG1) were significantly enriched in cardiac muscle cell myoblast differentiation (GO:0060379). CCL21 was one of the hub genes in the PPI network and also a target gene of miR-3144-5p. Moreover, the TF of v-Myc avian myelocytomatosis viral oncogene neuroblastoma-derived homolog (MYCN) was involved in the miR-3144-5p-TF-target gene coregulatory network and interacted with the target genes of miR-3144-5p.ISL1, NRG1, CCL21, and MYCN were differentially expressed in the miR-3144-5p mimic group, suggesting that miR-3144-5p overexpression plays a role in HCMs by regulating these genes and TF. This study may provide new insight into the mechanisms behind the progression of cardiac arrhythmia.
Project description:A precursor microRNA (miRNA) has two arms: miR-5p and miR-3p (miR-5p/-3p). Depending on the tissue or cell types, both arms can become functional. However, little is known about their coregulatory mechanisms during the tumorigenic process. Here, by using the large-scale miRNA expression profiles of five cancer types, we revealed that several of miR-5p/-3p arms were concordantly dysregulated in each cancer. To explore possible coregulatory mechanisms of concordantly dysregulated miR-5p/-3p pairs, we developed a robust computational framework and applied it to lung cancer data. The framework deciphers miR-5p/-3p coregulated protein interaction networks critical to lung cancer development. As a novel part in the method, we uniquely applied the second-order partial correlation to minimize false-positive regulations. Using 279 matched miRNA and mRNA expression profiles extracted from tumor and normal lung tissue samples, we identified 17 aberrantly expressed miR-5p/-3p pairs that potentially modulate the gene expression of 35 protein complexes. Functional analyses revealed that these complexes are associated with cancer-related biological processes, suggesting the oncogenic potential of the reported miR-5p/-3p pairs. Specifically, we revealed that the reduced expression of miR-145-5p/-3p pair potentially contributes to elevated expression of genes in the "FOXM1 transcription factor network" pathway, which may consequently lead to uncontrolled cell proliferation. Subsequently, the regulation of miR-145-5p/-3p in the FOXM1signaling pathway was validated by a cohort of 104 matched miRNA and protein (reverse-phase protein array) expression profiles in lung cancer. In summary, our computational framework provides a novel tool to study miR-5p/-3p coregulatory mechanisms in cancer and other diseases.
Project description:Myocardial infarction (MI) is a severe coronary artery disease and a leading cause of mortality and morbidity worldwide. However, the molecular mechanisms of MI have yet to be fully elucidated. In this study, we compiled MI-related genes, MI-related microRNAs (miRNAs) and known human transcription factors (TFs), and we then identified 1,232 feed-forward loops (FFLs) among these miRNAs, TFs and their co-regulated target genes through integrating target prediction. By merging these FFLs, the first miRNA and TF mediated regulatory network for MI was constructed, from which four regulators (SP1, ESR1, miR-21-5p and miR-155-5p) and three regulatory modules that might play crucial roles in MI were then identified. Furthermore, based on the miRNA and TF mediated regulatory network and literature survey, we proposed a pathway model for miR-21-5p, the miR-29 family and SP1 to demonstrate their potential co-regulatory mechanisms in cardiac fibrosis, apoptosis and angiogenesis. The majority of the regulatory relations in the model were confirmed by previous studies, which demonstrated the reliability and validity of this miRNA and TF mediated regulatory network. Our study will aid in deciphering the complex regulatory mechanisms involved in MI and provide putative therapeutic targets for MI.
Project description:Hypertrophic cardiomyopathy (HCM) is the most common genetic cardiovascular disease. Although some genes and miRNAs related with HCM have been studied, the molecular regulatory mechanisms between miRNAs and transcription factors (TFs) in HCM have not been systematically elucidated. In this study, we proposed a novel method for identifying dysregulated miRNA-TF feed-forward loops (FFLs) by integrating sample matched miRNA and gene expression profiles and experimentally verified interactions of TF-target gene and miRNA-target gene. We identified 316 dysregulated miRNA-TF FFLs in HCM, which were confirmed to be closely related with HCM from various perspectives. Subpathway enrichment analysis demonstrated that the method was outperformed by the existing method. Furthermore, we systematically analysed the global architecture and feature of gene regulation by miRNAs and TFs in HCM, and the FFL composed of hsa-miR-17-5p, FASN and STAT3 was inferred to play critical roles in HCM. Additionally, we identified two panels of biomarkers defined by three TFs (CEBPB, HIF1A, and STAT3) and four miRNAs (hsa-miR-155-5p, hsa-miR-17-5p, hsa-miR-20a-5p, and hsa-miR-181a-5p) in a discovery cohort of 126 samples, which could differentiate HCM patients from healthy controls with better performance. Our work provides HCM-related dysregulated miRNA-TF FFLs for further experimental study, and provides candidate biomarkers for HCM diagnosis and treatment.
Project description:The present study was aimed to unravel the inhibitory mechanisms of curcumin for lung cancer metastasis via constructing a miRNA-transcription factor (TF)-target gene network. Differentially expressed miRNAs between human high-metastatic non-small cell lung cancer 95D cells treated with and without curcumin were identified using a TaqMan human miRNA array followed by real-time PCR, out of which, the top 6 miRNAs (miR-302b-3p, miR-335-5p, miR-338-3p, miR-34c-5p, miR-29c-3p and miR-34a-35p) with more verified target genes and TFs than other miRNAs as confirmed by a literature review were selected for further analysis. The miRecords database was utilized to predict the target genes of these 6 miRNAs, TFs of which were identified based on the TRANSFAC database. The findings of the above procedure were used to construct a miRNA-TF-target gene network, among which miR-34a-5p, miR-34c-5p and miR-302b-3p seemed to regulate CCND1, WNT1 and MYC to be involved in Wnt signaling pathway through the LEF1 transcription factor. Therefore, we suggest miR-34a-5p/miR-34c-5p/miR-302b-3p -LEF1-CCND1/WNT1/MYC axis may be a crucial mechanism in inhibition of lung cancer metastasis by curcumin.
Project description:Both exfoliation glaucoma (XFG) and primary open-angle glaucoma (POAG) have been linked to decreased conventional outflow of aqueous humor (AH). To better understand the molecular changes in the AH content under such conditions, we analyzed the miRNA profiles of AH samples from patients with POAG and XFG compared to non-glaucoma controls. Individual AH samples (n?=?76) were collected from POAG and XFG patients and age-matched controls during surgical procedure. After RNA extraction, the miRNA profiles were individually determined in 12 POAG, 12 XFG and 11 control samples. We identified 205, 295 and 195 miRNAs in the POAG, XFG and control samples, respectively. Our differential expression analysis identified three miRNAs (miR-125b-5p, miR-302d-3p and miR-451a) significantly different between POAG and controls, five miRNAs (miR-122-5p, miR-3144-3p, miR-320a, miR-320e and miR-630) between XFG and controls and one miRNA (miR-302d-3p) between POAG and XFG. While none of these miRNAs have been previously linked to glaucoma, miR-122-5p may target three glaucoma-associated genes: OPTN, TMCO1 and TGF-ß1. Pathway analysis revealed that these miRNAs are involved in potential glaucoma pathways, including focal adhesion, tight junctions, and TGF-ß signaling. Comparison of the miRNA profile in AH to unrelated human serum (n?=?12) exposed potential relationships between these two fluids, although they were not significantly correlated. In summary, we have successfully profiled the miRNA expression without amplification in individual human AH samples and identified several POAG or XFG-associated miRNAs. These miRNAs may play a role in pathways previously implicated in glaucoma and act as biomarkers for disease pathogenesis.
Project description:<b>Background: </b>Untreated nephropathy can progress to renal failure. The traditional Mongolian remedy Narenmandula regulates the kidney "yang." This study aimed to identify key microRNAs (miRNAs) targeted by Narenmandula in a rat model of nephropathy.<br><br><b>Methods: </b>Fifteen rats exhibiting normal renal function were randomized to three study arms. Nephropathy was induced in <i>n</i>?=?10 rats using doxorubicin hydrochloride, followed by either Narenmandula treatment (treatment group) or no treatment (control group). In <i>n</i>?=?5 rats, no doxorubicin was given and renal function remained unchanged (healthy group). Microarray analysis identified miRNAs which were differentially expressed (DE-miRNAs) between groups. Target genes of DE-miRNAs were predicted using miRWalk version 2.0, followed by enrichment analysis using DAVID, and construction of the miRNA coregulatory network using Cytoscape.<br><br><b>Results: </b>Nephropathy was successfully induced, with doxorubicin resulting in differential expression of 3645 miRNAs (1324 upregulated and 2321 downregulated). Narenmandula treatment induced differential expression of a total of 159 miRNAs (102 upregulated and 57 downregulated). Upregulated DE-miRNAs (e.g., miR-497-5p, miR-195-5p, miR-181a-5p, miR-181c-5p, and miR-30e-5p) and downregulated DE-miRNAs (e.g., miR-330-3p and miR-214-3p) regulated a high number of target genes. Moreover, the miRNA pairs (e.g., miR-195-5p-miR-497-5p, miR-181a-5p-miR-181c-5p, and miR-30e-5p-miR-30a-5p) coregulated a high number of genes. Enrichment analysis indicated functional synergy between miR-30e-5p-miR-30a-3p, miR-34a-5p-miR-30e-5p, miR-30e-5p-miR-195-3p, and miR-30a-3p-miR-195-3p pairs.<br><br><b>Conclusion: </b>Narenmandula may modulate doxorubicin-induced nephropathy via targeting miR-497-5p, miR-195-5p, miR-181a-5p, miR-181c-5p, miR-30e-5p, miR-330-3p, miR-214-3p, miR-34a-5p, miR-30a-3p, and miR-30a-5p.
Project description:Growing evidence indicates that microRNAs (miRNAs) have a key role in processes involved in type 1 diabetes mellitus (T1DM) pathogenesis, including immune system functions and beta-cell metabolism and death. Although dysregulated miRNA profiles have been identified in T1DM patients, results are inconclusive; with only few miRNAs being consistently dysregulated among studies. Thus, we performed a systematic review of the literature on the subject, followed by bioinformatic analysis, to point out which miRNAs are dysregulated in T1DM-related tissues and in which pathways they act. PubMed and EMBASE were searched to identify all studies that compared miRNA expressions between T1DM patients and non-diabetic controls. Search was completed in August, 2017. Those miRNAs consistently dysregulated in T1DM-related tissues were submitted to bioinformatic analysis, using six databases of miRNA-target gene interactions to retrieve their putative targets and identify potentially affected pathways under their regulation. Thirty-three studies were included in the systematic review: 19 of them reported miRNA expressions in human samples, 13 in murine models and one in both human and murine samples. Among 278 dysregulated miRNAs reported in these studies, 25.9% were reported in at least 2 studies; however, only 48 of them were analyzed in tissues directly related to T1DM pathogenesis (serum/plasma, pancreas and peripheral blood mononuclear cells (PBMCs)). Regarding circulating miRNAs, 11 were consistently dysregulated in T1DM patients compared to controls: miR-21-5p, miR-24-3p, miR-100-5p, miR-146a-5p, miR-148a-3p, miR-150-5p, miR-181a-5p, miR-210-5p, miR-342-3p, miR-375 and miR-1275. The bioinformatic analysis retrieved a total of 5867 validated and 2979 predicted miRNA-target interactions for human miRNAs. In functional enrichment analysis of miRNA target genes, 77 KEGG terms were enriched for more than one miRNA. These miRNAs are involved in pathways related to immune system function, cell survival, cell proliferation and insulin biosynthesis and secretion. In conclusion, eleven circulating miRNAs seem to be dysregulated in T1DM patients in different studies, being potential circulating biomarkers of this disease.
Project description:BackgroundLung adenocarcinoma (LUAD) is one of the most common cancers worldwide. The etiology and pathophysiology of LUAD remain unclear. The aim of the present study was to identify the key genes, miRNAs and transcription factors (TFs) associated with the pathogenesis and prognosis of LUAD.MethodsThree gene expression profiles (GSE43458, GSE32863, GSE74706) of LUAD were obtained from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) were identified by GEO2R.The Gene Ontology (GO) terms, pathways, and protein-protein interactions (PPIs) of these DEGs were analyzed. Bases on DEGs, the miRNAs and TFs were predicted. Furthermore, TF-gene-miRNA co-expression network was constructed to identify key genes, miRNAs and TFs by bioinformatic methods. The expressions and prognostic values of key genes, miRNAs and TFs were carried out through The Cancer Genome Atlas (TCGA) database and Kaplan Meier-plotter (KM) online dataset.ResultsA total of 337 overlapped DEGs (75 upregulated and 262 downregulated) of LUAD were identified from the three GSE datasets. Moreover, 851 miRNAs and 29 TFs were identified to be associated with these DEGs. In total, 10 hub genes, 10 key miRNAs and 10 key TFs were located in the central hub of the TF-gene-miRNA co-expression network, and validated using The Cancer Genome Atlas (TCGA) database. Specifically, seven genes (PHACTR2, MSRB3, GHR, PLSCR4, EPB41L2, NPNT, FBXO32), two miRNAs (hsa-let-7e-5p, hsa-miR-17-5p) and four TFs (STAT6, E2F1, ETS1, JUN) were identified to be associated with prognosis of LUAD, which have significantly different expressions between LUAD and normal lung tissue. Additionally, the miRNA/gene co-expression analysis also revealed that hsa-miR-17-5p and PLSCR4 have a significant negative co-expression relationship (r=?0.33, P=1.67e-14) in LUAD.ConclusionsOur study constructed a regulatory network of TF-gene-miRNA in LUAD, which may provide new insights about the interaction between genes, miRNAs and TFs in the pathogenesis of LUAD, and identify potential biomarkers or therapeutic targets for LUAD.
Project description:The SCN5A gene encodes cardiac sodium channel Nav1.5 and causes lethal ventricular arrhythmias/sudden death and atrial fibrillation (AF) when mutated. MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression, and involved in the pathogenesis of many diseases. However, little is known about the regulation of SCN5A by miRNAs. Here we reveal a novel post-transcriptional regulatory mechanism for expression and function of SCN5A/Nav1.5 via miR-192-5p. Bioinformatic analysis revealed that the 3'-UTR of human and rhesus SCN5A, but not elephant, pig, rabbit, mouse, and rat SCN5A, contained a target binding site for miR-192-5p and dual luciferase reporter assays showed that the site was critical for down-regulation of human SCN5A. With Western blot assays and electrophysiological studies, we demonstrated that miR-192-5p significantly reduced expression of SCN5A and Nav1.5 as well as peak sodium current density INa generated by Nav1.5. Notably, in situ hybridization, immunohistochemistry and real-time qPCR analyses showed that miR-192-5p was up-regulated in tissue samples from AF patients, which was associated with down-regulation of SCN5A/Nav1.5. These results demonstrate an important post-transcriptional role of miR-192-5p in post-transcriptional regulation of Nav1.5, reveal a novel role of miR-192-5p in cardiac physiology and disease, and provide a new target for novel miRNA-based antiarrhythmic therapy for diseases with reduced INa.
Project description:Calcium deposition in vascular smooth muscle cells (VSMCs) is a form of ectopic ossification in blood vessels. It can result in rigidity of the vasculature and an increase in cardiac events. Here, we report that the microRNA miR-134-5p potentiates inorganic phosphate (Pi)-induced calcium deposition in VSMCs by inhibiting histone deacetylase 5 (HDAC5). Using miRNA microarray analysis of Pi-treated rat VSMCs, we first selected miR-134-5p for further evaluation. Quantitative RT-PCR confirmed that miR-134-5p was increased in Pi-treated A10 cells, a rat VSMC line. Transfection of miR-134-5p mimic potentiated the Pi-induced increase in calcium contents. miR-134-5p increased the amounts of bone runt-related transcription factor 2 (RUNX2) protein and bone morphogenic protein 2 (BMP2) mRNA in the presence of Pi but decreased the expression of osteoprotegerin (OPG). Bioinformatic analysis showed that the HDAC5 3'untranslated region (3'UTR) was one of the targets of miR-134-5p. The luciferase construct containing the 3'UTR of HDAC5 was down-regulated by miR-134-5p mimic in a dose-dependent manner in VSMCs. Overexpression of HDAC5 mitigated the calcium deposition induced by miR-134-5p. Our results suggest that a Pi-induced increase of miR-134-5p may cause vascular calcification through repression of HDAC5.