miRNA-mediated Networks Research Articles

MiR-200/183 family-mediated module biomarker for gastric cancer progression: an AI-assisted bioinformatics method with experimental functional survey

BackgroundGastric cancer (GC) is a major cancer burden throughout the world with a high mortality rate. The performance of current predictive and prognostic factors is still limited. Integrated analysis is required for accurate cancer progression predictive biomarker and prognostic biomarkers that help to guide therapy.MethodsAn AI-assisted bioinformatics method that combines transcriptomic data and microRNA regulations were used to identify a key miRNA-mediated network module in GC progression. To reveal the module’s function, we performed the gene expression analysis in 20 clinical samples by qRT-PCR, prognosis analysis by multi-variable Cox regression model, progression prediction by support vector machine, and in vitro studies to elaborate the roles in GC cells migration and invasion.ResultsA robust microRNA regulated network module was identified to characterize GC progression, which consisted of seven miR-200/183 family members, five mRNAs and two long non-coding RNAs H19 and CLLU1. Their expression patterns and expression correlation patterns were consistent in public dataset and our cohort. Our findings suggest a two-fold biological potential of the module: GC patients with high-risk score exhibited a poor prognosis (p-value < 0.05) and the model achieved AUCs of 0.90 to predict GC progression in our cohort. In vitro cellular analyses shown that the module could influence the invasion and migration of GC cells.ConclusionsOur strategy which combines AI-assisted bioinformatics method with experimental and clinical validation suggested that the miR-200/183 family-mediated network module as a “pluripotent module”, which could be potential marker for GC progression.

Integrated transcriptome and small RNA sequencing in revealing miRNA-mediated regulatory network of floral bud break in Prunus mume.

MicroRNAs is one class of small non-coding RNAs that play important roles in plant growth and development. Though miRNAs and their target genes have been widely studied in many plant species, their functional roles in floral bud break and dormancy release in woody perennials is still unclear. In this study, we applied transcriptome and small RNA sequencing together to systematically explore the transcriptional and post-transcriptional regulation of floral bud break in P. mume. Through expression profiling, we identified a few candidate genes and miRNAs during different developmental stage transitions. In total, we characterized 1,553 DEGs associated with endodormancy release and 2,084 DEGs associated with bud flush. Additionally, we identified 48 known miRNAs and 53 novel miRNAs targeting genes enriched in biological processes such as floral organ morphogenesis and hormone signaling transudation. We further validated the regulatory relationship between differentially expressed miRNAs and their target genes combining computational prediction, degradome sequencing, and expression pattern analysis. Finally, we integrated weighted gene co-expression analysis and constructed miRNA-mRNA regulatory networks mediating floral bud flushing competency. In general, our study revealed the miRNA-mediated networks in modulating floral bud break in P. mume. The findings will contribute to the comprehensive understanding of miRNA-mediated regulatory mechanism governing floral bud break and dormancy cycling in wood perennials.

Integrated mRNA and Small RNA Sequencing Reveals a microRNA Regulatory Network Associated with Starch Biosynthesis in Lotus (Nelumbo nucifera Gaertn.) Rhizomes.

Internode starch biosynthesis is one of the most important traits in lotus rhizome because of its relation to crop productivity. Understanding the microRNA (miRNA) and mRNA expression profiles related to lotus internode starch biosynthesis would help develop molecular improvement strategies, but they are not yet well-investigated. To identify genes and miRNAs involved in internode starch biosynthesis, the cDNA and small RNA libraries of Z6-1, Z6-2, and Z6-3 were sequenced, and their expression were further studied. Through combined analyses of transcriptome data and small RNA sequencing data, a complex co-expression regulatory network was constructed, in which 20 miRNAs could modulate starch biosynthesis in different internodes by tuning the expression of 10 target genes. QRT-PCR analysis, transient co-expression experiment and dual luciferase assay comprehensively confirmed that NnumiR396a down-regulated the expression of NnSS2 and ultimately prevents the synthesis of amylopectin, and NnumiR396b down-regulated the expression of NnPGM2 and ultimately prevents the synthesis of total starch. Our results suggest that miRNAs play a critical role in starch biosynthesis in lotus rhizome, and that miRNA-mediated networks could modulate starch biosynthesis in this tissue. These results have provided important insights into the molecular mechanism of starch biosynthesis in developing lotus rhizome.

MiR-125-chinmo pathway regulates dietary restriction-dependent enhancement of lifespan in Drosophila.

Dietary restriction (DR) extends healthy lifespan in diverse species. Age and nutrient-related changes in the abundance of microRNAs (miRNAs) and their processing factors have been linked to organismal longevity. However, the mechanisms by which they modulate lifespan and the tissue-specific role of miRNA-mediated networks in DR-dependent enhancement of lifespan remains largely unexplored. We show that two neuronally enriched and highly conserved microRNAs, miR-125 and let-7 mediate the DR response in Drosophila melanogaster. Functional characterization of miR-125 demonstrates its role in neurons while its target chinmo acts both in neurons and the fat body to modulate fat metabolism and longevity. Proteomic analysis revealed that Chinmo exerts its DR effects by regulating the expression of FATP, CG2017, CG9577, CG17554, CG5009, CG8778, CG9527, and FASN1. Our findings identify miR-125 as a conserved effector of the DR pathway and open the avenue for this small RNA molecule and its downstream effectors to be considered as potential drug candidates for the treatment of late-onset diseases and biomarkers for healthy aging in humans.

MicroRNA transcriptomic analysis of the sixth leaf of maize (Zea mays L.) revealed a regulatory mechanism of jointing stage heterosis

BackgroundZhengdan 958 (Zheng 58 × Chang 7–2), a commercial hybrid that is produced in a large area in China, is the result of the successful use of the heterotic pattern of Reid × Tang-SPT. The jointing stage of maize is the key period from vegetative to reproductive growth, which determines development at later stages and heterosis to a certain degree. MicroRNAs (miRNAs) play vital roles in the regulation of plant development, but how they function in the sixth leaf at the six-leaf (V6) stage to influence jointing stage heterosis is still unclear.ResultOur objective was to study miRNAs in four hybrid combinations developed in accordance with the Reid × Tang-SPT pattern, Zhengdan 958, Anyu 5 (Ye 478 × Chang 7–2), Ye 478 × Huangzaosi, Zheng 58 × Huangzaosi, and their parental inbred lines to explore the mechanism related to heterosis. A total of 234 miRNAs were identified in the sixth leaf at the V6 stage, and 85 miRNAs were differentially expressed between the hybrid combinations and their parental inbred lines. Most of the differentially expressed miRNAs were non-additively expressed, which indicates that miRNAs may participate in heterosis at the jointing stage. miR164, miR1432 and miR528 families were repressed in the four hybrid combinations, and some miRNAs, such as miR156, miR399, and miR395 families, exhibited different expression trends in different hybrid combinations, which may result in varying effects on the heterosis regulatory mechanism.ConclusionsThe potential targets of the identified miRNAs are related to photosynthesis, the response to plant hormones, and nutrient use. Different hybrid combinations employ different mature miRNAs of the same miRNA family and exhibit different expression trends that may result in enhanced or repressed gene expression to regulate heterosis. Taken together, our results reveal a miRNA-mediated network that plays a key role in jointing stage heterosis via posttranscriptional regulation.

Identification of the Potential Prognostic Markers from the miRNA-lncRNA-mRNA Interactions for Metastatic Renal Cancer via Next-Generation Sequencing and Bioinformatics.

The survival rate in patients with metastatic renal cell carcinoma (RCC) is low. In addition, metastatic RCC resists traditional treatment. Therefore, identification of novel biomarkers, signaling pathways, and therapeutic targets is an important issue. The aim of the present study is to identify novel prognostic markers from the miRNA-mediated network for the regulation of metastasis of RCC. To address this issue, the RNA of human RCC cell lines, 786-O and ACHN, derived from primary and metastatic sites, respectively, were collected and subjected to RNA sequencing and small RNA sequencing. The bioinformatic analysis revealed that the pathways of the genes with different expressions were related to tumor progression, and identified miRNA and miRNA-long non-coding RNA (lncRNA) interactions, and mRNA. The results revealed that the expressions of seven miRNAs were associated with the overall survival rate of patients with RCC. Furthermore, the expressions of two lncRNA and three protein-coding genes (mRNA) were significantly associated with the increased or decreased disease-free survival rate. Although the detailed regulatory mechanism between miRNAs and targeted genes was not fully understood, our findings present novel prognostic markers and novel insight on miRNA-mediated pathways for metastatic RCC.

Identification of microRNAs in developing wheat grain that are potentially involved in regulating grain characteristics and the response to nitrogen levels

BackgroundMicroRNAs (miRNAs) play crucial roles in the regulation of plant development and growth, but little information is available concerning their roles during grain development under different nitrogen (N) application levels. Our objective was to identify miRNAs related to the regulation of grain characteristics and the response to different N fertilizer conditions.ResultsA total of 79 miRNAs (46 known and 33 novel miRNAs) were identified that showed significant differential expression during grain development under both high nitrogen (HN) and low nitrogen (LN) treatments. The miRNAs that were significantly upregulated early in grain development target genes involved mainly in cell differentiation, auxin-activated signaling, and transcription, which may be associated with grain size; miRNAs abundant in the middle and later stages target genes mainly involved in carbohydrate and nitrogen metabolism, transport, and kinase activity and may be associated with grain filling. Additionally, we identified 50 miRNAs (22 known and 28 novel miRNAs), of which 11, 9, and 39 were differentially expressed between the HN and LN libraries at 7, 17, and 27 days after anthesis (DAA). The miRNAs that were differentially expressed in response to nitrogen conditions target genes involved mainly in carbohydrate and nitrogen metabolism, the defense response, and transport as well as genes that encode ubiquitin ligase. Only one novel miRNA (PC-5p-2614_215) was significantly upregulated in response to LN treatment at all three stages, and 21 miRNAs showed significant differential expression between HN and LN conditions only at 27 DAA. We therefore propose a model for target gene regulation by miRNAs during grain development with N-responsive patterns.ConclusionsThe potential targets of the identified miRNAs are related to various biological processes, such as carbohydrate/nitrogen metabolism, transcription, cellular differentiation, transport, and defense. Our results indicate that miRNA-mediated networks, via posttranscriptional regulation, play crucial roles in grain development and the N response, which determine wheat grain weight and quality. Our study provides useful information for future research of regulatory mechanisms that focus on improving grain yield and quality.

Integrated transcriptome, small RNA, and degradome analysis reveals the complex network regulating starch biosynthesis in maize

BackgroundStarch biosynthesis in endosperm is a key process influencing grain yield and quality in maize. Although a number of starch biosynthetic genes have been well characterized, the mechanisms by which the expression of these genes is regulated, especially in regard to microRNAs (miRNAs), remain largely unclear.ResultsSequence data for small RNAs, degradome, and transcriptome of maize endosperm at 15 and 25 d after pollination (DAP) from inbred lines Mo17 and Ji419, which exhibit distinct starch content and starch granule structure, revealed the mediation of starch biosynthetic pathways by miRNAs. Transcriptome analysis of these two lines indicated that 33 of 40 starch biosynthetic genes were differentially expressed, of which 12 were up-regulated in Ji419 at 15 DAP, one was up-regulated in Ji419 at 25 DAP, 14 were up-regulated in Ji419 at both 15 and 25 DAP, one was down-regulated in Ji419 at 15 DAP, two were down-regulated in Ji419 at 25 DAP, and three were up-regulated in Ji419 at 15 DAP and down-regulated in Ji419 at 25 DAP, compared with Mo17. Through combined analyses of small RNA and degradome sequences, 22 differentially expressed miRNAs were identified, including 14 known and eight previously unknown miRNAs that could target 35 genes. Furthermore, a complex co-expression regulatory network was constructed, in which 19 miRNAs could modulate starch biosynthesis in endosperm by tuning the expression of 19 target genes. Moreover, the potential operation of four miRNA-mediated pathways involving transcription factors, miR169a-NF-YA1-GBSSI/SSIIIa and miR169o-GATA9-SSIIIa/SBEIIb, was validated via analyses of expression pattern, transient transformation assays, and transactivation assays.ConclusionOur results suggest that miRNAs play a critical role in starch biosynthesis in endosperm, and that miRNA-mediated networks could modulate starch biosynthesis in this tissue. These results have provided important insights into the molecular mechanism of starch biosynthesis in developing maize endosperm.

Identification of Differentially Expressed miRNAs between a Wheat K-type Cytoplasmic Male Sterility Line and Its Near-Isogenic Restorer Line.

K-type cytoplasmic male sterility (KCMS) lines were ideal material for three-line hybrid wheat system due to the major role in hybrid wheat production. In this study, the morphology of developing microspore and mature pollen was compared between a KCMS line and its near-isogenic restorer line (KCMS-NIL). The most striking difference is that the microspore was unable to develop into tricellular pollen in the KCMS line. MicroRNA plays vital roles in flowering and gametophyte development. Small RNA sequencing identified a total of 274 known and 401 novel miRNAs differentially expressed between two lines or two developmental stages. Most of miRNAs with high abundance were differentially expressed at the uninucleate stage, and their expression level recovered or remained at the binucleate stage. Further degradome sequencing identified target genes which were mainly enriched in transcription regulation, phytohormone signaling and RNA degradation pathways. Combining with the transcriptome data, a correlation was found between the abnormal anther development, such as postmeiotic mitosis cessation, deformative pollen wall and the chromosome condensation of the vegetative cell, and the alterations in the related miRNA and their targets expression profiles. According to the correlation and pathway analysis, we propose a hypothetic miRNA-mediated network for the control of KCMS restoration.

DPMIND: degradome-based plant miRNA-target interaction and network database.

MicroRNAs (miRNAs) play essential roles in plant growth, development and stress responses through post-transcriptionally regulating the expression levels of their target mRNAs. Although some tools and databases were developed for predicting the relationships between miRNAs and their targets (miR-Tar), most of them were dependent on computational methods without experimental validations. With the development of degradome sequencing techniques, researchers can identify potential interactions based on degradome sequencing data. The validations with specific degradome data are useful to identify the miR-Tar interactions (MTIs) occurring in/under some specific tissues or treatments. Degradome-based plant miRNA-target interaction and network database (DPMIND) collected almost all available plant degradome data and built a retrieval and analysis platform of miRNA-target interactions and miRNA regulatory networks (MRNs). DPMIND contains the recently updated 3794 miRNAs and 28666 verified MTIs with 69 degradomes from 10 plant species. Not only the verified MTIs but also the degradome-based MRNs can be retrieved from DPMIND. Users can search for the verified MTIs and build degradome-based MRNs for the specific miRNAs or targets. DPMIND can build the MRNs based on all degradomes or specific degradomes, which helps to identify all possible connections among specific miR-Tar and compare the miRNA-mediated networks among various tissues or treatments. It can also build the networks mediated by all known miRNAs based on specific degradomes. Furthermore, DPMIND can be used to study the conservation and specificity of MTIs and sub-networks across different plant tissues or species. http://202.195.246.60/DPMIND/. huangji@njau.edu.cn. Supplementary data are available at Bioinformatics online.

‘Traffic light rules’: Chromatin states direct miRNA-mediated network motifs running by integrating epigenome and regulatome

BackgroundEpigenetic marks can cooperatively regulate chromatin accessibility and in turn facilitate or impede the binding of regulatory factors to various elements, suggesting their important roles in regulatory circuits. However, it remains elusive as to how epigenetic marks cooperate in the operations of regulatory network. MethodsHere, we systematically characterized chromatin states of 26 epigenetic marks on different elements of protein-coding genes and miRNAs. We comprehensively analyzed, by using an integrative regulatory network, how cooperation among epigenetic, transcriptional, and post-transcriptional regulations came about. ResultsWe observed extensive cooperation of epigenetic marks on local functional elements and complex epigenetic patterns corresponding to different biological functions. By identifying the significantly epigenetic state-modified motifs, we found that multiple combinations of epigenetic states were associated with a specific type of motif. Interestingly, miRNA-mediated motifs were linked to stable epigenetic states of downstream targets. Changes in epigenetic states of downstream targets in miRNA-mediated motifs can buffer the effects of upstream regulator on target genes, suggesting that miRNA-mediated motifs require the cooperation of epigenetic marks. ConclusionsOverall, epigenetic marks are involved in the running of regulatory motifs in the way traffic lights control traffic flows and hence should be part of the architecture of complex regulatory circuits. General significanceWe demonstrated a detailed analysis of the cooperation of multiple epigenetic marks and how epigenetic regulation was organized into a human regulatory network. The findings form a basis for further understanding of the complicated roles of epigenetic marks on regulatory circuits.

Integrated analyses to reconstruct microRNA-mediated regulatory networks in mouse liver using high-throughput profiling.

BackgroundMicroRNAs (miRNAs) simultaneously target many transcripts through partial complementarity binding, and have emerged as a key type of post-transcriptional regulator for gene expression. How miRNA accomplishes its pleiotropic effects largely depends on its expression and its target repertoire. Previous studies discovered thousands of miRNAs and numerous miRNA target genes mainly through computation and prediction methods which produced high rates of false positive prediction. The development of Argonaute cross-linked immunoprecipitation coupled with high-throughput sequencing (CLIP-Seq) provides a system to effectively determine miRNA target genes. Likewise, the accuracy of dissecting the transcriptional regulation of miRNA genes has been greatly improved by chromatin immunoprecipitation of the transcription factors coupled with sequencing (ChIP-Seq). Elucidation of the miRNA target repertoire will provide an in-depth understanding of the functional roles of microRNA pathways. To reliably reconstruct a miRNA-mediated regulatory network, we established a computational framework using publicly available, sequence-based transcription factor-miRNA databases, including ChIPBase and TransmiR for the TF-miRNA interactions, along with miRNA-target databases, including miRTarBase, TarBase and starBase, for the miRNA-target interactions. We applied the computational framework to elucidate the miRNA-mediated regulatory network in the Mir122a-/- mouse model, which has an altered transcriptome and progressive liver disease.ResultsWe applied our computational framework to the expression profiles of miRNA/mRNA of Mir122a-/- mutant mice and wild-type mice. The miRNA-mediated network involves 40 curated TFs contributing to the aberrant expression of 65 miRNAs and 723 curated miRNA target genes, of which 56% was found in the differentially-expressed genes of Mir122a--mice. Hence, the regulatory network disclosed previously-known and also many previously-unidentified miRNA-mediated regulations in mutant mice. Moreover, we demonstrate that loss of imprinting at the chromosome 12qF1 region is associated with miRNA overexpression in human hepatocellular carcinoma and stem cells, suggesting initiation of precancerous changes in young mice deficient in miR-122. A group of 9 miRNAs was found to share miR-122 target genes, indicating synergy between miRNAs and target genes by way of multiplicity and cooperativity.ConclusionsThe study provides significant insight into miRNA-mediated regulatory networks. Based on experimentally verified data, this network is highly reliable and effective in revealing previously-undetermined disease-associated molecular mechanisms. This computational framework can be applied to explore the significant TF-miRNA-miRNA target interactions in any complex biological systems with high degrees of confidence.

MicroRNA-mediated networks underlie immune response regulation in papillary thyroid carcinoma

Papillary thyroid carcinoma (PTC) is a common endocrine malignancy with low death rate but increased incidence and recurrence in recent years. MicroRNAs (miRNAs) are small non-coding RNAs with diverse regulatory capacities in eukaryotes and have been frequently implied in human cancer. Despite current progress, however, a panoramic overview concerning miRNA regulatory networks in PTC is still lacking. Here, we analyzed the expression datasets of PTC from The Cancer Genome Atlas (TCGA) Data Portal and demonstrate for the first time that immune responses are significantly enriched and under specific regulation in the direct miRNA−target network among distinctive PTC variants to different extents. Additionally, considering the unconventional properties of miRNAs, we explore the protein-coding competing endogenous RNA (ceRNA) and the modulatory networks in PTC and unexpectedly disclose concerted regulation of immune responses from these networks. Interestingly, miRNAs from these conventional and unconventional networks share general similarities and differences but tend to be disparate as regulatory activities increase, coordinately tuning the immune responses that in part account for PTC tumor biology. Together, our systematic results uncover the intensive regulation of immune responses underlain by miRNA-mediated networks in PTC, opening up new avenues in the management of thyroid cancer.

Toward microRNA-mediated gene regulatory networks in plants

Current achievements in plant microRNA (miRNA) research area are inspiring. Molecular cloning and functional elucidation have greatly advanced our understanding of this small RNA species. As one of the ultimate goals, many research efforts devoted to draw a comprehensive view of miRNA-mediated gene regulatory networks in plants. Numerous bioinformatics tools competent for network analysis have been available. However, the most important point for network construction is to obtain reliable analytical results based on sufficient experimental data. Here, we introduced a general workflow to retrieve and analyze the desired data sets that serve as the cornerstones for network construction. For the upstream analyses of miRNA genes, the sequence feature of miRNA promoters should be characterized. And, regulatory relationships between transcription factors (TFs) and miRNA genes need to be investigated. For the downstream part, we emphasized that the high-throughput degradome sequencing data were especially useful for genuine miRNA-target pair identification. Functional characterization of the miRNA targets is essential to provide deep biological insights into certain miRNA-mediated pathways. For miRNAs themselves, studies on their organ- or tissue-specific expression patterns and the mechanism of self-regulation were discussed. Besides, exhaustive literature mining is required to further support or improve the established networks. It is desired that the introduced framework for miRNA-mediated network construction is timely and useful and could inspire more research efforts in the miRNA research area.

New insight into the role of miRNAs in leukemia

Recent studies have shown that microRNAs(miRNAs) play an important role in cell differentiation, growth, and death, including the functional study of miRNAs in tumorigenesis. To date, miRNA expression profiles in many types of cancers have been identified and miRNA expression signatures associated with types and cytogenetics of leukemia have also been reported. Increasing evidence has shown that miRNAs could function as either tumor suppressors or oncogenes in cancers such as leukemia, while other miRNAs might be benefitcial for diagnosis and prognosis, predicted to be newly developed biomarkers. In this review, we summarize the recent progress about miRNAs in leukemia and present a miRNA-mediated network involved in differentiation, proliferation and apoptosis predicted to be the roles of miRNAs in the pathogenesis of leukemia.