Role Of miRNAs In Response Research Articles

Temporal expression analysis of microRNAs and their target GRAS genes induced by osmotic stress in two contrasting wheat genotypes.

MicroRNAs (miRNAs) are important nonprotein-coding genes in plants which participate in almost all biological processes during abiotic and biotic stresses. Understanding how plants respond to various environmental conditions requires the identification of stress-related miRNAs. In recent years, there has been an increased interest in studying miRNA genes and gene expression. Drought is one of the common environmental stresses limiting plant growth and development. Stress-specific miRNAs and their GRAS gene targets were validated to understand the role of miRNAs in response to osmotic stress. In this study, expression patterns of the ten stress-responsive miRNAs involved in osmotic stress adaptation were examined in order to undertand the regulation behavior of abiotic stress and miRNAs in two contrasting wheat genotype C-306 (drought tolerant) and WL-711 (drought sensitive). Three miRNAs were discovered to be upregulated under stress, whereas seven miRNAs were showed to be down-regulated as a consequence of the study. In contrast to miRNA, it was also discovered that GRAS genes as their targets were up-regulated during osmotic stress. In addition, the expression level of miR159, miR408 along with their targets, TaGRAS178 and TaGRAS84 increased in response to osmotic stress. Nevertheless, miR408 is highly conserved miRNA that regulates plant growth, development and stress response. As a result, variation in the expression levels of studied miRNAs in the presence of target genes provides a plausible explanation for miRNA-based abiotic stress regulation. A regulatory network of miRNA and their targets revealed that fourteen miRNA interact with 55 GRAS targets from various subfamilies that contribute in the plant growth and development. These findings provide evidence for temporal and variety-specific differential regulation of miRNAs and their targets in wheat in response to osmotic shock, and they may aid in determining the potential.

Small RNA Profiling Reveals a Role of miRNAs in Response to Ralstonia solanacearum Infection in Tomato

Small RNA Profiling Reveals a Role of miRNAs in Response to Ralstonia solanacearum Infection in Tomato

Genome-wide post-transcriptional regulation of bovine mammary gland response to Streptococcus uberis.

MicroRNAs (miRNAs) as post-transcriptionally regulators of gene expression have been shown to be critical regulators to fine-tuning immune responses, besides their criteria for being an ideal biomarker. The regulatory role of miRNAs in responses to most mastitis-causing pathogens is not well understood. Gram-positive Streptococcus uberis (Str. uberis), the leading pathogen in dairy herds, cause both clinical and subclinical infections. In this study, a system biology approach was used to better understand the main post-transcriptional regulatory functions and elements of bovine mammary gland response to Str. uberis infection. Publicly available miRNA-Seq data containing 50 milk samples of the ten dairy cows (five controls and five infected) were retrieved for this current research. Functional enrichment analysis of predicted targets revealed that highly confident responsive miRNAs (4 up- and 19 downregulated) mainly regulate genes involved in the regulation of transcription, apoptotic process, regulation of cell adhesion, and pro-inflammatory signaling pathways. Time series analysis showed that six gene clusters significantly differed in comparisons between Str. uberis-induced samples with controls. Additionally, other bioinformatic analysis, including upstream network analysis, showed essential genes, including TP53 and TGFB1 and some small molecules, including glucose, curcumin, and LPS, commonly regulate most of the downregulated miRNAs. Upregulated miRNAs are commonly controlled by the most important genes, including IL1B, NEAT1, DICER1 enzyme and small molecules including estradiol, tamoxifen, estrogen, LPS, and epigallocatechin. Our study used results of next-generation sequencing to reveal key miRNAs as the main regulator of gene expression responses to a Gram-positive bacterial infection. Furthermore, by gene regulatory network (GRN) analysis, we can introduce the common upregulator transcription factor of these miRNAs. Such milk-based miRNA signature(s) would facilitate risk stratification for large-scale prevention programs and provide an opportunity for early diagnosis and therapeutic intervention.

Integrated Analyses of miRNome and Transcriptome Reveal the Critical Role of miRNAs Toward Heat Stress Response in Isochrysis galbana.

Isochrysis galbana is widely used in aquaculture as a bait microalgal species. High temperature (HT) can severely impair the development of I. galbana, exerting adverse effects on its yield. MicroRNAs (miRNAs) play an essential role in modulating stress-responsive genes. However, the role of miRNAs in response to HT in microalgae remains largely unexplored. In the present study, we identified several conserved and novel miRNAs in I. galbana through miRNome sequencing. Among these identified miRNAs, 22 miRNAs were differentially expressed in response to heat stress, and their target genes were predicted accordingly. Moreover, a comprehensive and integrated analysis of miRNome and transcriptome was performed. We found that six potential reversely correlated differentially expressed miRNA (DEM) and differentially expressed gene (DEG) pairs were associated with heat stress response (HSR) in I. galbana. The expressions of DEMs and DEGs were further verified using real-time quantitative PCR (RT-qPCR). Integrated analyses showed that miRNAs played fundamental roles in the regulatory network of HSR in I. galbana mainly by regulating some heat-responsive genes, including heat shock proteins (HSPs), reactive oxygen species (ROS) signaling-related genes, and specific key genes in the ubiquitination pathway. Our current study identified the first set of heat-responsive miRNAs from I. galbana and helped elucidate the miRNA-mediated HSR and resistance mechanisms in I. galbana. This new knowledge could provide ways to enhance its heat stress tolerance.

An Integration of MicroRNA and Transcriptome Sequencing Analysis Reveal Regulatory Roles of miRNAs in Response to Chilling Stress in Wild Rice.

A chromosome single segment substitution line (CSSL) DC90, which was generated by introgressing CTS-12, a locus derived from common wild rice (Oryza rufipogon Griff.), into the 9311 (Oryza sativa L. ssp. indica) background, exhibits a chilling tolerance phenotype under chilling stress. Here, an integration of microRNA (miRNA) deep sequencing and transcriptomic sequencing analysis was performed to explore the expression profiles of miRNAs and their target genes mediated by CTS-12 under chilling stress, and to reveal the possible regulatory mechanisms of miRNAs that are involved in chilling tolerance. Integration analysis revealed that a number of differentially expressed miRNAs (DEMs) and putative target genes with different expression patterns and levels were identified in 9311 and DC90 under chilling stress. KEGG enrichment analysis revealed that the target genes that are regulated by chilling-induced miRNAs are involved in the regulation of various biological processes/pathways, including protein biosynthesis, redox process, photosynthetic process, and chloroplast development in two genotypes. CRISPR/Cas9 editing of the target genes of the key DEMs in a chilling tolerant rice variety Zhonghua 11 (ZH11) found that LOC_Os11g48020 (OsGL1-11), one of the putative target genes of osa-miR1846a/b-5p and encoding a wax synthesis protein, is correlated with a chilling stress tolerance phenotype, implying osa-miR1846a/b-5p/OsGL1-11 plays an important role in CTS-12-mediated chilling stress tolerance regulatory pathway(s). Therefore, we speculate that the CTS-12 may regulate the key miRNA target genes in response to chilling stress by differential regulation of miRNAs in wild rice, thereby resulting in the variation of chilling tolerance phenotype between 9311 and DC90.

Epigenetic Regulation of Cardiac Troponin Genes in Pediatric Patients with Heart Failure Supported by Ventricular Assist Device.

Ventricular Assist Device (VAD) therapy is considered as a part of standard care for end-stage Heart Failure (HF) children unresponsive to medical management, but the potential role of miRNAs in response to VAD therapy on molecular pathways underlying LV remodeling and cardiac function in HF is unknown. The aims of this study were to evaluate the effects of VAD on miRNA expression profile in cardiac tissue obtained from HF children, to determine the putative miRNA targets by an in-silico analysis as well as to verify the changes of predicated miRNA target in the same cardiac samples. The regulatory role of selected miRNAs on predicted targets was evaluated by a dedicated in vitro study. miRNA profile was determined in cardiac samples obtained from 13 HF children [median: 29 months; 19 LVEF%; 9 Kg] by NGS before VAD implant (pre-VAD) and at the moment of heart transplant (Post-VAD). Only hsa-miR-199b-5p, hsa-miR-19a-3p, hsa-miR-1246 were differentially expressed at post-VAD when compared to pre-VAD, and validated by real-time PCR. Putative targets of the selected miRNAs were involved in regulation of sarcomere genes, such as cardiac troponin (cTns) complex. The expression levels of fetal ad adult isoforms of cTns resulted significantly higher after VAD in cardiac tissue of HF pediatric patients when compared with HF adults. An in vitro study confirmed a down-regulatory effect of hsa-miR-19a-3p on cTnC expression. The effect of VAD on sarcomere organization through cTn isoform expression may be epigenetically regulated, suggesting for miRNAs a potential role as therapeutic targets to improve heart function in HF pediatric patients.

Small RNA analysis of Perna viridis after exposure to Prorocentrum lima, a DSP toxins-producing dinoflagellate.

Diarrheic shellfish poisoning toxins (DSP toxins) are a set of the most important phycotoxins produced by some dinoflagellates. Studies have shown that DSP toxins have various toxicities such as genotoxicity, cytotoxicity, and immunotoxicity to bivalve mollusks. However, these toxicities appear decreasing with exposure time and concentration of DSP toxins. The underlying mechanism involved remains unclear. In this study, small RNA sequencing was performed in the digestive gland of the mussel Perna viridis after exposure to DSP toxins-producing dinoflagellate Prorocentrum lima for different time periods. The potential roles of miRNAs in response and detoxification to DSP toxins in the mussel were analyzed. Small RNA sequencing of 12 samples from 72 individuals was conducted by BGISEQ-500. A total of 123 mature miRNAs were identified, including 90 conserved miRNAs and 33 potential novel miRNAs. After exposure to P. lima, multiple important miRNAs displayed some alterations. Further miRNA target prediction revealed some important genes involved in cytoskeleton, apoptosis, complement system and immune stress. qPCR demonstrated that miR-71_5, miR-750_1 and novel_mir4 were significantly up-regulated at 6 h after exposure to P. lima, while miR-100_2 was significantly down-regulated after 96 h of exposure. Accordingly, putative target genes of these differentially expressed miRNAs experienced some changes. After 6 h of DSP toxins exposure, NHLRC2 and C1q-like were significantly down-regulated. After 96 h of DSP toxins exposure, NHLRC2 was significantly up-regulated. It is reasonable to speculate that the mussel P. viridis might respond to DSP toxins through miR-750_1, novel_mir4 and miR-71_5 regulating the expression of relevant target genes involved in apoptosis, cytoskeleton, and immune response, etc. This study might provide new clues to uncover the toxic response of bivalve to DSP toxins and lay a foundation for revealing the roles of miRNAs in the environmental adaptation in shellfish.

The Effect of Low Temperature Stress on the Leaves and MicroRNA Expression of Potato Seedlings

In recent years, with the wanton destruction of the ecological environment by humans and the frequent occurrence of extreme bad weather, many places that should have been warm and blooming in spring have instead experienced the phenomenon of the “April blizzard,” which has seriously affected China's crops, especially spring potato production in most areas. Potato cultivars, especially potato seedlings, are sensitive to frost, and low temperature frost has become one of the most important abiotic stresses affecting potato production. Potato cold tolerance is regulated by a complex gene network. Although some low temperature resistant microRNAs have been identified, little is known about the role of miRNAs in response to low temperature stress in potato. Therefore, the objective of this study is to clarify the influence of low temperature stress on the miRNA expression of potato by comparing the expression differences of miRNA in potato which was treated with different low temperatures. For the study, 307 known miRNAs belonging to 73 small RNA families and 211 novel miRNAs were obtained. When the temperature decreased, the number of both known and novel miRNA decreased, and the minimum temperature was −2°C. Most of the miRNAs respond to low temperature, drought, and disease stress; some conserved miRNAs were first found to respond to low temperature stress in potato, such as stu-miR530, stu-miR156d, and stu-miR167b. The Gene Ontology, Kyoto Encyclopedia of Genes, and Genomes pathway enrichment analysis of 442 different expression miRNAs target genes indicated that there existed diversified low temperature responsive pathways, but Abscisic Acid was found likely to play a central coordinating role in response to low temperature stress in many metabolism pathways. Quantitative real-time PCR assays indicated that the related targets were negatively regulated by the tested different expression miRNAs during low temperature stress. The results indicated that miRNAs may play an important coordination role in response to low temperature stress in many metabolic pathways by regulating abscisic acid and gibberellin, which provided insight into the roles of miRNAs during low temperature stress and would be helpful for alleviating low temperature stress and promoting low temperature resistant breeding in potatoes.

Molecular insight into regulation of miRNAs in the spleen of zebrafish (Danio rerio) upon pathogenic Streptococcus parauberis infection

MicroRNAs (miRNAs) constitute a group of small non-coding RNAs (~22 nucleotides) and one of their main functions is to regulate the immune responses. Gram-positive bacterium, Streptococcus parauberis is the main causative agent of “Streptococcosis” in wide range of fish species. In this study, we performed high throughput sequencing analysis to identify the miRNA profile against S. parauberis infection in the spleen of zebrafish (Danio rerio). Overall, 349 known and 151 novel miRNAs were discovered. Among them, 12 known miRNAs (dre-miR-34b, dre-miR-135a, dre-miR-200b-5p, dre-miR-146b, dre-miR-31, dre-miR-17a-3p, dre-miR-222a-3p, dre-miR-731, dre-miR-301b-3p and dre-miR-30a-3p) and 9 novel miRNAs were differentially expressed (DE) in the spleen of S. parauberis challenged zebrafish. The identified 12 DE miRNAs were predicted to regulate 721 target genes. We confirmed the miRNA expression results by validating selected known and novel DE miRNAs using qRT-PCR. Gene Ontology (GO), Kyoto Encyclopedia of Genes (KEGG) pathway analysis and miRNA-mRNA interactions implies that specific target genes of DE miRNAs are associated with immune responses. The enriched pathways included Toll-like receptor (TLR), C-type lectin, NOD-like receptor, and RIG-I-like receptor signaling pathways, etc. Especially, dre-miR-200b-5p, dre-miR-146b, dre-miR-731, dre-miR-222a-3p, and dre-miR-34b were able to target potential immune-related genes such as il10, irak1, traf6, hspa8 and ikbke upon S. parauberis challenge. Thus, overall results could lay a foundation to understand the underlying immune regulatory role of miRNAs in response to pathogenic S. parauberis infection in teleosts.

Integrative analysis of microRNA and mRNA expression profiles in MARC-145 cells infected with PRRSV

Porcine reproductive and respiratory syndrome virus (PRRSV) causes tremendous economic losses to the swine industry worldwide. miRNAs are crucial regulators of gene expression and a wide range of complex interactions of miRNAs-mRNAs is possible during virus infection. However, there is no comprehensive integrated study of miRNA and mRNA networks in MARC-145 cells after infection with PRRSV. We analyzed the differential expressions, co-relations, annotations, and putative functions of miRNA and mRNA networks in PRRSV-infected MARC-145 cells. Based on the filtering criterion, 22 differentially expressed miRNAs (DEmiRs) (15 up- and 7 downregulated) were filtered out. miRNA-mRNA interaction networks were constructed. For the 18 selected miRNAs, 390 potential target genes were predicted from the differentially expressed mRNAs (DEmRs). GO and KEGG pathway annotations predicted 34 KEGG pathways, 12 of which are known to be involved in virus infection. Real-time PCR validated the RNA-seq results. Our analysis showed that miR-27a-5p and miR-21-3p downregulate the expression of two of their potential target genes-SPARC, CLIC1, and cofilin-1, COX7A2, respectively. Further experiments proved that miR-21-3p and miR-27a-5p can promote PRRSV replication significantly. It is the first report that these two miRNAs participate in the interaction of host cells with PRRSV. Our results provide insights into the role of miRNAs in response to PRRSV infection, which will aid the research for developing novel therapies against PRRSV.

Integrated small RNA and mRNA expression profiles reveal miRNAs and their target genes in response to Aspergillus flavus growth in peanut seeds

BackgroundMicroRNAs are important gene expression regulators in plants immune system. Aspergillus flavus is the most common causal agents of aflatoxin contamination in peanuts, but information on the function of miRNA in peanut-A. flavus interaction is lacking. In this study, the resistant cultivar (GT-C20) and susceptible cultivar (Tifrunner) were used to investigate regulatory roles of miRNAs in response to A. flavus growth.ResultsA total of 30 miRNAs, 447 genes and 21 potential miRNA/mRNA pairs were differentially expressed significantly when treated with A. flavus. A total of 62 miRNAs, 451 genes and 44 potential miRNA/mRNA pairs exhibited differential expression profiles between two peanut varieties. Gene Ontology (GO) analysis showed that metabolic-process related GO terms were enriched. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses further supported the GO results, in which many enriched pathways were related with biosynthesis and metabolism, such as biosynthesis of secondary metabolites and metabolic pathways. Correlation analysis of small RNA, transcriptome and degradome indicated that miR156/SPL pairs might regulate the accumulation of flavonoids in resistant and susceptible genotypes. The miR482/2118 family might regulate NBS-LRR gene which had the higher expression level in resistant genotype. These results provided useful information for further understanding the roles of miR156/157/SPL and miR482/2118/NBS-LRR pairs.ConclusionsIntegration analysis of the transcriptome, miRNAome and degradome of resistant and susceptible peanut varieties were performed in this study. The knowledge gained will help to understand the roles of miRNAs of peanut in response to A. flavus.

Identification of Regulatory Networks of MicroRNAs and Their Targets in Response to Colletotrichum gloeosporioides in Tea Plant (Camellia sinensis L.).

Anthracnose disease is caused by Colletotrichum gloeosporioides, and is common in leaves of the tea plant (Camellia sinensis). MicroRNAs (miRNAs) have been known as key modulators of gene expression in response to environmental stresses, disease resistance, defense responses, and plant immunity. However, the role of miRNAs in responses to C. gloeosporioides remains unexplored in tea plant. Therefore, in the present study, six miRNA sequencing data sets and two degradome data sets were generated from C. gloeosporioides-inoculated and control tea leaves. A total of 485 conserved and 761 novel miRNAs were identified. Of those, 239 known and 369 novel miRNAs exhibited significantly differential expression under C. gloeosporioides stress. One thousand one hundred thirty-four and 596 mRNAs were identified as targets of 389 conserved and 299 novel miRNAs by degradome analysis, respectively. Based on degradome analysis, most of the predicted targets are negatively correlated with their corresponding conserved and novel miRNAs. The expression levels of 12 miRNAs and their targets were validated by quantitative real-time PCR. A negative correlation between expression profiles of five miRNAs (PC-5p-80764_22, csn-miR160c, csn-miR828a, csn-miR164a, and csn-miR169e) and their targets (WRKY, ARF, MYB75, NAC, and NFY transcription factor) was observed. The predicted targets of five interesting miRNAs were further validated through 5’RLM-RACE. Furthermore, Gene Ontology and metabolism pathway analysis revealed that most of the target genes were involved in the regulation of auxin pathway, ROS scavenging pathway, salicylic acid mediated pathway, receptor kinases, and transcription factors for plant growth and development as well as stress responses in tea plant against C. gloeosporioides stress. This study enriches the resources of stress-responsive miRNAs and their targets in C. sinensis and thus provides novel insights into the miRNA-mediated regulatory mechanisms, which could contribute to the enhanced susceptibility of C. gloeosporioides in tea plant.

MicroRNAs and new biotechnological tools for its modulation and improving stress tolerance in plants.

SummaryMicroRNAs (miRNAs) modulate the abundance and spatial–temporal accumulation of target mRNAs and indirectly regulate several plant processes. Transcriptional regulation of the genes encoding miRNAs (MIR genes) can be activated by numerous transcription factors, which themselves are regulated by other miRNAs. Fine‐tuning of MIR genes or miRNAs is a powerful biotechnological strategy to improve tolerance to abiotic or biotic stresses in crops of economic importance. Current approaches for miRNA fine‐tuning are based on the down‐ or up‐regulation of MIR gene transcription and the use of genetic engineering tools to manipulate the final concentration of these miRNAs in the cytoplasm. Transgenesis, cisgenesis, intragenesis, artificial MIR genes, endogenous and artificial target mimicry, MIR genes editing using Meganucleases, ZNF proteins, TALENs and CRISPR/Cas9 or CRISPR/Cpf1, CRISPR/dCas9 or dCpf1, CRISPR13a, topical delivery of miRNAs and epigenetic memory have been successfully explored to MIR gene or miRNA modulation and improve agronomic traits in several model or crop plants. However, advantages and drawbacks of each of these new biotechnological tools (NBTs) are still not well understood. In this review, we provide a brief overview of the biogenesis and role of miRNAs in response to abiotic or biotic stresses, we present critically the main NBTs used for the manipulation of MIR genes and miRNAs, we show current efforts and findings with the MIR genes and miRNAs modulation in plants, and we summarize the advantages and drawbacks of these NBTs and provide some alternatives to overcome. Finally, challenges and future perspectives to miRNA modulating in important crops are also discussed.

MicroRNAs and response to therapy in leukemia

A variety of epigenetic factors involved in leukemia pathogenesis. Among various epigenetic factors, microRNAs (miRNAs) have emerged as important players, which affect a sequence of cellular and molecular signaling pathways. Leukemia is known as progressive cancer, which is related to many health problems in the world. It has been shown that the destruction of the blood-forming organs could lead to abnormal effects on the proliferation and development of leukocytes and their precursors. Despite many attempts for approved effective and powerful therapies for patients with leukemia, finding and developing new therapeutic approaches are required. One of the important aspects of leukemia therapy, identification of underlying cellular and molecular mechanisms involved in the pathogenesis of leukemia. Several miRNAs (ie, miR-103, miR-101, mit-7, let-7i, miR-424, miR-27a, and miR-29c) and play major roles in response to therapy in patients with leukemia. miRNAs exert their effects by targeting a variety of targets, which are associated with response to therapy in patients with leukemia. It seems that more understanding about the roles of miRNAs in response to therapy in patients with leukemia could contribute to better treatment of patients with leukemia. Here, for the first time, we summarized various miRNAs, which are involved in response to therapy in the treatment patients with leukemia.

Characterization of cadmium-responsive MicroRNAs and their target genes in maize (Zea mays) roots

BackgroundCurrent research has shown that microRNAs (miRNAs) play vital roles in plant response to stress caused by heavy metals such as aluminum, arsenic, cadmium (Cd), and mercury. Cd has become one of the most hazardous pollutants in the environment. Maize can be a potential model to study phytoremediation of Cd-contaminated soil owing to its large biomass production. However, little is known about miRNAs as a response to Cd stress in maize.ResultsTo investigate the role of miRNAs in response to Cd stress, roots of seedlings of the inbred maize lines B73 and Mo17 were collected and treated with 200 mg/L CdCl2·2.5 H2O over different exposure times. Enzyme activities of superoxide dismutase and peroxidase were measured to confirm Cd stress. The expression of six candidate miRNAs and their targets were validated using quantitative real-time PCR (qRT-PCR) technology. In addition, the expression of Zma-miR171b was assessed using in situ hybridization.ConclusionsOur results showed that miRNAs and their respective target genes were differentially expressed in maize seedling roots exposed to Cd stress. This research produced new insights into the molecular mechanism of miRNAs responsive to Cd stress in plants and sheds light on the latent roles of miRNAs in plants exposed to heavy metal stresses.

Integrating miRNA and mRNA expression profiles in plasma of laying hens associated with heat stress.

High temperature is one of the most common environmental stressors plaguing animal husbandry worldwide. Little is known about the regulatory roles of miRNAs in response to heat stress in laying hens. To systematically identify heat stress-responsive miRNAs and their targets in laying hens, the differential expression of miRNAs and mRNAs was compared under heat stress and normal temperature. We identified 16 miRNAs and 502 genes that were significantly changed in heat-stressed laying hens. By comparing the differentially expressed genes (DEGs) and the putative targets of the altered miRNAs based on bioinformatics prediction, 82 coordinated genes were identified. Gene ontology classification analyses of the 82 putative target genes showed that the biological category 'cellular response to stress' was prominently annotated. Notably, the response-related gene autophagy-related protein 9A was most likely controlled by the upregulated miRNAs gga-miR-92-5p, gga-miR-1618-5p, gga-miR-1737, and gga-miR-6557 in response to heat stress. Analysis of DEGs also revealed an increase in lipid metabolism in heat-stressed laying hens. Some of these genes were negatively correlated with the altered miRNAs, suggesting that they are potential targets of the miRNAs. Taken together, our results advance our understanding of the regulatory mechanism of heat-stress-induced injury in laying hens, specifically with regard to miRNAs.

Novel microRNAs are associated with population divergence in transcriptional response to thermal stress in an intertidal copepod.

The role of gene expression in adaptation to differing thermal environments has been assayed extensively. Yet, in most natural systems, analyses of gene expression reveal only one level of the complexity of regulatory machineries. MicroRNAs (miRNAs) are small noncoding RNAs which are key components of many gene regulatory networks, and they play important roles in a variety of cellular pathways by modulating post-transcriptional quantities of mRNA available for protein synthesis. The characterization of miRNA loci and their regulatory dynamics in nonmodel systems are still largely understudied. In this study, we examine the role of miRNAs in response to high thermal stress in the intertidal copepod Tigriopus californicus. Allopatric populations of this species show varying levels of local adaptation with respect to thermal regimes, and previous studies showed divergence in gene expression between populations from very different thermal environments. We examined the transcriptional response to temperature stress in two populations separated by only 8km by utilizing RNA-seq to quantify both mRNA and miRNA levels. Using the currently available genome sequence, we first describe the repertoire of miRNAs in T.californicus and assess the degree to which transcriptional response to temperature stress is governed by miRNA activity. The two populations showed large differences in the number of genes involved, the magnitude of change in commonly used genes and in the number of miRNAs involved in transcriptional modulation during stress. Our results suggest that an increased level of regulatory network complexity may underlie improved survivorship under thermal stress in one of the populations.

Identification and characterization of drought responsive microRNAs and their target genes in cardamom (Elettaria cardamomum Maton)

Plant miRNAs are found to be present throughout the genome and they regulate gene expression either by cleaving mRNA or inhibiting the translational process at the post transcriptional level. Drought is one of the major limiting factors that negatively affect productivity of plants. Cardamom cultivation is having good production potential but the plants are vulnerable to biotic and abiotic stress factors. To date, nothing is known about the regulatory roles of miRNAs in response to drought stress in cardamom. Ion Torrent sequencing of two small RNA libraries prepared from control (C) and treated (T) plants raised under well irrigated and drought stressed treatments respectively created 3,938,342 and 4,083,181 primary reads. A total of 150 conserved and 20 novel microRNAs were identified from both the control and treated libraries. Discovery of 17 differentially expressed miRNAs under drought stress suggests that these miRNAs might have involved in various biological processes to improve plant tolerance to water stress. Several target genes for drought stress regulating miRNAs were identified including miR156l and miR169c which cleave the target mRNA involved in response to water deprivation. miR530b and miR156a target mRNAs which respond to water deprivation and inhibit the translational process. The expression patterns of some of the miRNAs and their targets were validated by qRT-PCR. This study is the first report of drought responsive miRNAs and their targets in cardamom. The outcome of this research could provide insights into the miRNA based regulatory mechanisms in response to drought stress in monocot plants.

Regulation through MicroRNAs in Response to Low-Energy N+ Ion Irradiation in Oryza sativa.

MicroRNAs (miRNAs) are a non-coding regulatory RNAs that play significant roles in plant growth and development, especially in the stress response. Low-energy ion radiation, a type of environmental stress, can cause multiple biological effects. To understand the roles of miRNAs in response to low-energy N+ ion radiation in Oryza sativa, high-throughput sequencing of small RNAs was carried out to detect the expression of miRNAs in the shoots of the rice after 2 × 1017 N+/cm2 irradiation. The differentially expressed 28 known miRNAs were identified, 17 of these identified miRNAs were validated by real-time quantitative fluorescent PCR (q-PCR), including 9 up-regulated miRNAs (miR1320-3p, miR1320-5p, miR156b-3p, miR156c-5p, miR156c-3p/g-3p, miR1561-5p, miR398b and miR6250) and 8 down-regulated miRNAs (miR156a/e/i, miR156k, miR160f-5p, miR166j-5p, miR1846e and miR399d). In addition, 45 novel radiationresponsive miRNAs were predicted, and 8 of them were verified by q-PCR. The target genes of radiation-responsive miRNAs were predicted and gene function enrichment analysis was performed with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). The expression of 9 targets of 4 known miRNA families (miR156, miR399, miR1320 and miR398) and 2 targets of 2 novel miRNAs were quantified by q-PCR, and a strong negative regulation relation between miRNAs and their targets were observed. Those targets including SQUAMOSA promoterbinding-like protein (SPL) genes, copper/zinc superoxide dismutase (Cu/Zn-SOD), copper chaperone for SOD (CCS1) and electron transporter/ heat-shock protein binding protein (HSP), which are involved in growth and defense against various stresses, especially associated with reactive oxygen species (ROS) scavenging. This work provides important information for understanding the ROS generation and elimination mechanisms closely related to miRNAs in rice seedlings after low-energy N+ radiation exposure.

High-throughput sequencing of small RNAs revealed the diversified cold-responsive pathways during cold stress in the wild banana (Musa itinerans)

BackgroundCold stress is one of the most severe abiotic stresses affecting the banana production. Although some miRNAs have been identified, little is known about the role of miRNAs in response to cold stress in banana, and up to date, there is no report about the role of miRNAs in the response to cold stress in the plants of the cultivated or wild bananas.ResultHere, a cold-resistant line wild banana (Musa itinerans) from China was used to profile the cold-responsive miRNAs by RNA-seq during cold stress. Totally, 265 known mature miRNAs and 41 novel miRNAs were obtained. Cluster analysis of differentially expressed (DE) miRNAs indicated that some miRNAs were specific for chilling or 0 °C treated responses, and most of them were reported to be cold-responsive; however, some were seldom reported to be cold-responsive in response to cold stress, e.g., miR395, miR408, miR172, suggesting that they maybe play key roles in response to cold stress. The GO and KEGG pathway enrichment analysis of DE miRNAs targets indicated that there existed diversified cold-responsive pathways, and miR172 was found likely to play a central coordinating role in response to cold stress, especially in the regulation of CK2 and the circadian rhythm. Finally, qPCR assays indicated the related targets were negatively regulated by the tested DE miRNAs during cold stress in the wild banana.ConclusionsIn this study, the profiling of miRNAs by RNA-seq in response to cold stress in the plants of the wild banana (Musa itinerans) was reported for the first time. The results showed that there existed diversified cold-responsive pathways, which provided insight into the roles of miRNAs during cold stress, and would be helpful for alleviating cold stress and cold-resistant breeding in bananas.