microRNA-mediated Regulation Research Articles

MiR-21-3p Centric Regulatory Network in Dairy Cow Mammary Epithelial Cell Proliferation.

MicroRNA-mediated gene regulation is important for the development of the mammary gland and the lactating process. A previous study has shown that the expression of microRNA-21 (miR-21) is different in the dry and early lactation period of the dairy cow mammary gland, but the molecular mechanisms underlying the lactation cycle are not fully understood. Here, the function of miR-21-3p on bovine mammary gland epithelial cells (BMECs) was detected by MTT assay and flow cytometry analysis, which showed that miR-21-3p significantly promoted the cell viability and proliferation. Then, the regulating mechanism of miR-21-3p on cell viability and proliferation was elucidated. Dual luciferase assay, RT-qPCR, and Western blot results revealed that IGFBP5 was a target gene of miR-21-3p. It was known that lncRNA could act as a competing endogenous RNA to sequester miRNAs and reduce the regulatory effect of miRNA-targeted genes. Based on our previous lncRNA-seq data and bioinformatics analysis, lncRNA NONBTAT017009.2 was potentially associated with miR-21-3p, and its expression was specifically inhibited with the transfection of miR-21-3p mimic into BMECs. Inversely, the overexpression of NONBTAT017009.2 significantly decreased the expression level of miR-21-3p in BMECs, while the expression of IGFBP5, the target gene of miR-21-3p, was significantly upregulated. In addition, the promoter region of miR-21 contained two STAT3 binding sites, and the dual luciferase reporter assays revealed that the overexpression of STAT3 significantly reduced the promoter activity of miR-21, implying that the transcription factor STAT3 may act as an upstream regulator affecting the regulation process of miR-21-3p. The overexpression of STAT3 significantly inhibited the expression of miR-21-3p, while the mRNA expression of IGFBP5 was significantly increased compared with the control group. Besides, there are no STAT3 binding sites in the promoter region of IGFBP5 as we predicted by gene-regulation and JASPAR software. Therefore, it could infer that STAT3 might regulate the expression of IGFBP5 by miR-21-3p. Taken together, these results established a regulatory network of miR-21-3p to illustrate the regulating mechanism on promoting cow mammary epithelial cell proliferation.

UPF1/SMG7-dependent microRNA-mediated gene regulation

The stability and quality of metazoan mRNAs are under microRNA (miRNA)-mediated and nonsense-mediated control. Although UPF1, a core mediator of nonsense-mediated mRNA decay (NMD), mediates the decay of target mRNA in a 3′UTR-length-dependent manner, the detailed mechanism remains unclear. Here, we suggest that 3′UTR-length-dependent mRNA decay is not mediated by nonsense mRNAs but rather by miRNAs that downregulate target mRNAs via Ago-associated UPF1/SMG7. Global analyses of mRNAs in response to UPF1 RNA interference in miRNA-deficient cells reveal that 3′UTR-length-dependent mRNA decay by UPF1 requires canonical miRNA targeting. The destabilization of miRNA targets is accomplished by the combination of Ago2 and UPF1/SMG7, which may recruit the CCR4-NOT deadenylase complex. Indeed, loss of the SMG7-deadenylase complex interaction increases the levels of transcripts regulated by UPF1-SMG7. This UPF1/SMG7-dependent miRNA-mediated mRNA decay pathway may enable miRNA targeting to become more predictable and expand the miRNA-mRNA regulatory network.

MiR-96 acts as a tumor suppressor via targeting the BCR-ABL1 oncogene in chronic myeloid leukemia blastic transformation

MicroRNA-mediated posttranscriptional regulation is an important epigenetic regulatory mechanism of gene expression, and its dysregulation is involved in the development and progression of a variety of malignancies, including chronic myeloid leukemia (CML). The BCR-ABL1 fusion gene is not only the initiating factor of CML, but it is also an important driving factor for blastic transformation. Tyrosine kinase inhibitors (TKIs) targeting BCR-ABL1 tyrosine kinase activity, represented by imatinib, are currently the first-line treatment for CML. However, due to primary resistance or secondary resistance caused by mutations in the BCR-ABL1 kinase domain, TKIs cannot completely prevent the progression of CML; thus, the study of BCR-ABL1 gene expression regulation is of great significance. In this study, bioinformatics analysis and our results showed that miR-96 could directly bind to the 3'UTR region of BCR-ABL1 to regulate fusion protein expression, thereby regulating its downstream signaling pathway activity. We also found that miR-96 was downregulated during the progression from the chronic phase (CML-CP) to the blast crisis (CML-BC). Downregulation of miR-96 could promote the proliferation and participate in the cell differentiation of CML-BC cells. Additionally, we found that the novel histone deacetylase drug chidamide and the DNA methyltransferase inhibitor decitabine could restore the low expression of miR-96 in CML cells, and there were two abnormal hypermethylated sites in the promoter region of miR-96 in CML, suggesting that its low expression might be at least partially regulated by epigenetic mechanisms. In addition, re-expression of miR-96 could increase the sensitivity of CML-BC cells to imatinib. Thus, miR-96 functions as a tumor suppressor, and re-expression of this microRNA might have therapeutic benefits in CML blastic transformation.

Genetic polymorphisms and epigenetic regulation of survivin encoding gene, BIRC5, in multiple sclerosis patients

BackgroundThe persistent the inflammatory condition in multiple sclerosis (MS) may due to the aberrant regulation of the elimination of the pathogenic autoreactive lymphocytes through apoptosis. Survivin, encoded by the BIRC5 gene, has been indicated to be involved in the regulation of apoptosis. This survey intended to investigate the genetic and microRNA mediated regulation of survivin in relapsing-remitting MS (RRMS) disease.ResultsIt was observed that the C allele (OR = 1.38, 95% CI = 1.05–1.348, P = 0.022) and CC genotype (OR = 1.84, 95% CI = 1.06–3.19; P = 0.029) in the rs9904341 polymorphism increased the disease risk. Furthermore, miR-34a was significantly downregulated (Fold change = 0.41, P = 0.001) in the PBMCs from RRMS subjects. Survivin mRNA expression in PBMCs and serum survivin level were increased in RRMS patients in comparison to the controls. Downregulation of miR-34a was negatively correlated with increased survivin level.ConclusionAlthough the genetic polymorphism of BIRC5 gene was associated with the disease risk, miR-34a was suggested to be involved in the regulation of survivin in the RRMS patients.

Unravelling the MicroRNA-Mediated Gene Regulation in Developing Pongamia Seeds by High-Throughput Small RNA Profiling.

Pongamia (Millettia pinnata syn. Pongamia pinnata) is a multipurpose biofuel tree which can withstand a variety of abiotic stresses. Commercial applications of Pongamia trees may substantially benefit from improvements in their oil-seed productivity, which is governed by complex regulatory mechanisms underlying seed development. MicroRNAs (miRNAs) are important molecular regulators of plant development, while relatively little is known about their roles in seed development, especially for woody plants. In this study, we identified 236 conserved miRNAs within 49 families and 143 novel miRNAs via deep sequencing of Pongamia seeds sampled at three developmental phases. For these miRNAs, 1327 target genes were computationally predicted. Furthermore, 115 differentially expressed miRNAs (DEmiRs) between successive developmental phases were sorted out. The DEmiR-targeted genes were preferentially enriched in the functional categories associated with DNA damage repair and photosynthesis. The combined analyses of expression profiles for DEmiRs and functional annotations for their target genes revealed the involvements of both conserved and novel miRNA-target modules in Pongamia seed development. Quantitative Real-Time PCR validated the expression changes of 15 DEmiRs as well as the opposite expression changes of six targets. These results provide valuable miRNA candidates for further functional characterization and breeding practice in Pongamia and other oilseed plants.

Integrated analysis of miRNAs and their targets reveals that miR319c/TCP2 regulates apical bud burst in tea plant (Camellia sinensis)

The roles of microRNA-mediated epigenetic regulation were highlighted in the bud dormancy-activity cycle, implying that certain differentially expressed miRNAs play crucial roles in apical bud burst, such as csn-miR319c/TCP2. microRNAs (miRNAs) are a class of small non-coding RNAs that regulate gene expression by targeting mRNA transcripts for cleavage or directing translational inhibition. To investigate whether miRNAs regulate bud dormancy-activation transition in tea plant, which largely affects the yield and price of tea products and adaptability of tea trees, we constructed small RNA libraries from three different periods of bud dormancy-burst transition. Through sequencing analysis, 262 conserved and 83 novel miRNAs were identified, including 118 differentially expressed miRNAs. Quantitative RT-PCR results for randomly selected miRNAs exhibited that our comprehensive analysis is highly reliable and accurate. The content of caffeine increased continuously from the endodormancy bud to flushing bud, and differentially expressed miRNAs coupling with their targets associated with bud burst were identified. Remarkably, csn-miR319c was downregulated significantly from the quiescent bud to burst bud, while its target gene CsnTCP2 (TEOSINTE BRANCHED/CYCLOIDEA/PROLIFERATING CELL FACTOR 2) displayed opposite expression patterns. Co-transformation experiment in tobacco demonstrated that csn-miR319c can significantly suppress the functions of CsnTCP2. This study on miRNAs and the recognition of target genes could provide new insights into the molecular mechanism of the bud dormancy-activation transition in tea plant.

Epigenetic mechanisms related to cognitive decline during aging.

Cognitive decline is a hallmark of the aging nervous system, characterized by increasing memory loss and a deterioration of mental capacity, which in turn creates a favorable context for the development of neurodegenerative diseases. One of the most detrimental alterations that occur at the molecular level in the brain during aging is the modification of the epigenetic mechanisms that control gene expression. As a result of these epigenetic-driven changes in the transcriptome most of the functions of the brain including synaptic plasticity, learning, and memory decline with aging. The epigenetic mechanisms altered during aging include DNA methylation, histone modifications, nucleosome remodeling, and microRNA-mediated gene regulation. In this review, we examine the current evidence concerning the changes of epigenetic modifications together with the molecular mechanisms underlying impaired neuronal gene transcription during aging. Herein, we discuss the alterations of DNA methylation pattern that occur in old neurons. We will also describe the most prominent age-related histone posttranslational modifications in the brain since changes in acetylation and methylation of specific lysine residues on H3 and H4 are associated to functional decline in the old. In addition, we discuss the role that changes in the levels of certain miRNAs would play in cognitive decline with aging. Finally, we provide an overview about the mechanisms either extrinsic or intrinsic that would trigger epigenetic changes in the aging brain, and the consequences of these changes, i.e., altered transcriptional profile and reactivation of transposable elements in old brain.

Therapeutic Delivery of miR-29b Enhances Radiosensitivity in Cervical Cancer.

Radioresistant cervical cancer is likely to give rise to local recurrence, distant metastatic relapse, and decreased survival rates. Recent studies revealed microRNA mediated regulation of tumor aggressiveness and metastasis; however, whether specific microRNAs regulate tumor radioresistance and can be exploited as radiosensitizing agents remains unclear. Here, we find that miR-29b could promote radiosensitivity in radioresistant subpopulations of cervical cancer cells. Notably, therapeutic delivery of miR-29b mimics via R11-SSPEI nanoparticle, whose specificity has been proved by our previous studies, can sensitize the tumor to radiation in a xenograft model. Mechanistically, we reveal a novel function of miR-29b in regulating intracellular reactive oxygen species signaling and explore a potential application for its use in combination with therapies known to increase oxidative stress such as radiation. Moreover, miR-29b inhibits DNA damage repair by targeting phosphate and tension homology deleted on chromsome ten (PTEN), and overexpression of PTEN could partially rescue miR-29b-mediated homologous recombination (HR)-DNA damage repair and increase radiosensitivity. These findings identify miR-29b as a radiosensitizing microRNA and reveal a new therapeutic strategy for radioresistant tumors.

Genome-wide analysis in Hevea brasiliensis laticifers revealed species-specific post-transcriptional regulations of several redox-related genes

MicroRNA-mediated post-transcriptional regulation has been reported on ROS production and scavenging systems. Although microRNAs first appeared highly conserved among plant species, several aspects of biogenesis, function and evolution of microRNAs were shown to differ. High throughput transcriptome and degradome analyses enable to identify small RNAs and their mRNA targets. A non-photosynthetic tissue particularly prone to redox reactions, laticifers from Hevea brasiliensis, revealed species-specific post-transcriptional regulations. This paper sets out to identify the 407 genes of the thirty main redox-related gene families harboured by the Hevea genome. There are 161 redox-related genes expressed in latex. Thirteen of these redox-related genes were targeted by 11 microRNAs. To our knowledge, this is the first report on a mutation in the miR398 binding site of the cytosolic CuZnSOD. A working model was proposed for transcriptional and post-transcriptional regulation with respect to the predicted subcellular localization of deduced proteins.

Development of the urogenital system is regulated via the 3\u2032UTR of GDNF

Mechanisms controlling ureter lenght and the position of the kidney are poorly understood. Glial cell-line derived neurotrophic factor (GDNF) induced RET signaling is critical for ureteric bud outgrowth, but the function of endogenous GDNF in further renal differentiation and urogenital system development remains discursive. Here we analyzed mice where 3′ untranslated region (UTR) of GDNF is replaced with sequence less responsive to microRNA-mediated regulation, leading to increased GDNF expression specifically in cells naturally transcribing Gdnf. We demonstrate that increased Gdnf leads to short ureters in kidneys located in an abnormally caudal position thus resembling human pelvic kidneys. High GDNF levels expand collecting ductal progenitors at the expense of ureteric trunk elongation and result in expanded tip and short trunk phenotype due to changes in cell cycle length and progenitor motility. MEK-inhibition rescues these defects suggesting that MAPK-activity mediates GDNF’s effects on progenitors. Moreover, Gdnf hyper mice are infertile likely due to effects of excess GDNF on distal ureter remodeling. Our findings suggest that dysregulation of GDNF levels, for example via alterations in 3′UTR, may account for a subset of congenital anomalies of the kidney and urinary tract (CAKUT) and/or congenital infertility cases in humans and pave way to future studies.

MicroRNA-mediated Regulation of Mucin-type O-glycosylation Pathway: A Putative Mechanism of Salivary Gland Dysfunction in Sjögren Syndrome.

To investigate microRNA (miRNA) that is potentially implicated in primary Sjögren syndrome (pSS)-related salivary hypofunction in labial salivary glands and to study miRNA-mediated mechanisms underlying oral dryness and altered rheology, focusing on the mucin O-glycosylation pathway. We performed miRNA expression profiling in minor salivary gland samples of patients with pSS presenting a different impairment in their unstimulated salivary flow rate. A computational in silico analysis was performed to identify genes and pathways that might be modulated by the deregulated miRNA that we had identified. To confirm in silico analysis, expression levels of genes encoding for glycosyltransferases and glycan-processing enzymes were investigated using Human Glycosylation-RT2 Profiler PCR Array. Among 754 miRNA analyzed, we identified 126 miRNA that were significantly deregulated in pSS compared to controls, with a trend that was inversely proportional with the impairment of salivary flow rates. An in silico approach pinpointed that several upregulated miRNA in patients with pSS target important genes in the mucin O-glycosylation. We confirmed this prediction by quantitative real-time PCR, highlighting the downregulation of some glycosyltransferase and glycosidase genes in pSS samples compared to controls, such as GALNT1, responsible for mucin-7 glycosylation. Collectively, our data suggest that the expression of different predicted miRNA-target genes in the mucin type O-glycan biosynthesis pathway is altered in pSS patients with low salivary flow and that the miRNA expression profile could influence the glycosidase expression levels and consequently the rheology in pSS.

MicroRNA-mediated regulation of BM-MSCs differentiation into sweat gland-like cells: targeting NF-κB.

Sweat gland regeneration is important for patients with an extensive deep burn injury. In previous study, we reported that bone marrow-mesenchymal stem cells (BM-MSCs) could differentiate into sweat gland-like cells (SGLCs), but the underlying molecular mechanism remains unclear. Recently, microRNAs(miRNAs or miRs) are reported to manipulate many biological processes. However, whether the process of MSCs differentiation into sweat gland cells (SGCs) is regulated by miRNAs has not been reported. In this study, BM-MSCs were induced into SGLCs by co-culturing with SGCs. Differential expressions of miRNAs between BM-MSC and SGLCs were determined through miRNAs microarray and 68 miRNAs were found significantly changed in miRNA profile including hsa-miR-138-5p. Bioinformatics analysis showed that hsa-miR-138-5p targeted a group of nuclear factor-κB (NF-κB) related genes which play an important role in skin appendage development. As expected, hsa-miR-138-5p inhibitor transfected into BM-MSCs partly mimicked the effects of co-culture and increased the number of SGLCs by increasing the expression of NF-κB related genes. These results suggest that hsa-miR-138-5p and NF-κB are involved in the regulation of BM-MSCs differentiation into SGLCs. This study may also offer a new approach to yield SGCs for burn patients.

MicroRNA-mediated redox regulation modulates therapy resistance in cancer cells: clinical perspectives.

Chemotherapy and radiation therapy are the most common types of cancer therapy. The development of chemo/radio-resistance remains, however, a major obstacle. Altered redox balances are among of the main factors mediating therapy resistance. Therefore, redox regulatory strategies are urgently needed to overcome this problem. Recently, microRNAs have been found to act as major redox regulatory factors affecting chemo/radio-resistance. MicroRNAs play critical roles in regulating therapeutic resistance through the regulation of antioxidant enzymes, redox-sensitive signaling pathways, cancer stem cells, DNA repair mechanisms and autophagy. Here, we summarize current knowledge on microRNA-mediated redox regulatory mechanisms underlying chemo/radio-resistance. This knowledge may form a basis for a better clinical management of cancer patients.

Nucleus-localized adiponectin is survival gatekeeper through miR-214-mediated AIFM2 regulation.

Adiponectin secreted from adipocytes into plasma has anti-aging, anti-obesity and anti-inflammation effects. Here, we detected intracellular adiponectin localized in the nuclei of human and mouse pluripotent stem cells, mouse germ cells and some somatic cells. Nucleus-localized (Nu) adiponectin protein is characterized by an N-terminal truncated monomer form in a native state, compared with intact multimer forms of cytoplasm-localized (Cy) adiponectin protein. Doxycycline-induced over-expression of ADIPONECTIN caused cell death in human and mouse Nu-type pluripotent stem cells. Genome-wide gene expression analysis indicated that apoptosis by ADIPONECTIN over-expression was induced in accompany with upregulation of AIFM2 and MEG3. Upregulation of AIFM2 and MEG3 and down-regulation of miR-214-3p verified by qPCR analyses after ADIPONECTIN over-expression indicated that the MEG3/miR-214/AIFM2 pathway played a role in the apoptotic cell death of pluripotent cells. Adiponectin-induced cell death was rescued by the treatment with miR-214-3p mimic. Global data analysis shows that Nu adiponectin has a role in microRNA-mediated post-transcription regulation, cell-cell interactions and chromatin remodeling as a survival gatekeeper.

SA94 - A DIRECT REGULATORY LINK BETWEEN MICRORNA MIR-137 AND SHANK2 WITH IMPLICATIONS FOR NEUROPSYCHIATRIC DISORDERS

Background The SHANK gene family encodes for postsynaptic scaffolding proteins with an important role at glutamatergic synapses in the brain and has been linked to a spectrum of neurodevelopmental disorders. The schizophrenia associated microRNA miR-137 and the SHANK genes converge on several levels, (a) both are expressed at the synapse, (b) both influence neuronal development, and (c) both have a strong link to neurodevelopmental and neuropsychiatric disorders like intellectual disability, autism and schizophrenia. This compiled evidence raised the question if the SHANKs might be targets of miR-137. Methods The in silico analysis of all three SHANK genes identified a single, highly conserved binding site for miR-137 in the 3´UTR of SHANK2. Luciferase assays were carried out in a neuroblastoma cell line (SH-SY5Y) and in mouse primary hippocampal neurons to validate the putative binding site. MiR-137 was overexpressed or inhibited in hippocampal neurons and Shank2 expression was analyzed by qPCR and Western blot. In addition, we analyzed expression levels of experimentally validated miR-137 target genes in the dorsolateral prefrontal cortex (DLPFC) of schizophrenia and control individuals using the RNA-seq data from the CommonMind Consortium. Results We can show that miR-137 targets the 3´UTR of SHANK2 in a site-specific manner. Overexpression of miR-137 significantly lowered endogenous Shank2 protein levels without detectable influence on mRNA level; conversely, the inhibition of miR-137 resulted in the increase of Shank2 protein in neuronal cells. To further support the link between miR-137 and schizophrenia, we compared the expression of mir-137 precursor and miR-137 target genes (including SHANK2) in the DLPFC of schizophrenia and control individuals. Almost one third (18/63; 29%) of validated miR-137 target genes showed significant expression differences in the DLPFC of schizophrenia individuals compared to controls and we propose that further targets (like SHANK2) may be regulated on protein level. Discussion In this study we identified SHANK2 as a novel direct target of miR-137. We discovered that physiological levels of miR-137 regulate SHANK2 expression and that the most likely mechanism of microRNA-mediated regulation is by repressing SHANK2 protein translation rather than mRNA degradation. Our study provides first evidence of a direct regulatory link between miR-137 and SHANK2 and supports the finding that miR-137 signaling is altered in schizophrenia.

Functional annotation of differentially expressed fetal cardiac microRNA targets: implication for microRNA-based cardiovascular therapeutics.

Gene expression pattern of a failing heart depicts remarkable similarity with developing fetal heart. Elucidating genetic as well as epigenetic mechanisms regulating the gene expression during cardiac development will improve our understanding of cardiovascular diseases. In the present study, we aimed to validate and characterize differentially expressed known microRNAs (miRNA) obtained from next generation sequencing data of two fetal cardiac developmental stages (days 4th and 14th) from chicken (G. gallus domesticus) using bioinformatic approaches. Potential mRNA targets of individual miRNA were identified and classified according to their biological, cellular, and molecular functions. Functional annotation of putative target genes was performed to predict their association with cardiovascular diseases. We identified a total of 19 differentially expressed miRNAs between 4th and 14th day sample from the data sets obtained by next generation sequencing. A total of nearly 1522 potential targets ranging from 15 to 270 for each miRNA were predicted out of which 1221 were unique, while 301 were overlapping. Gene ontology and KEGG analysis revealed that majority of these target genes regulate critical cellular and molecular processes including transcriptional regulation, protein transport, signal transduction, matrix remodeling, Ras signaling, MAPK signaling, and TGF-beta signaling pathways indicating the complex nature of microRNA-mediated gene regulation during cardiogenesis. We found a significant association between potential target genes and cardiovascular diseases validating a link between fetal cardiac miRNAs and regulation of cardiovascular disease-related genes. These important findings may lay a foundation for further understanding the regulatory mechanisms operative in gene re-programming in the failing heart.

Target-mimicry based diminution of miRNA167 reinforced flowering-time phenotypes in tobacco via spatial-transcriptional biases of flowering-associated miRNAs

Evolutionarily conserved microRNAs such as miR156, miR159, miR167 and miR172 tightly regulate the extensive array of gene expression during flowering in plants, through instant and long-term alterations in the expression of their target genes. Here we employed a novel target-mimicry approach for the diminution of auxin signalling regulator miRNA167 by developing mimic-transgenic lines in tobacco, to investigate the transcriptional biases of flowering-associated miRNAs in apical and floral meristematic tissues and their phenotypic implications. Recorded morpho-alterations such as uneven flowering-time phenotypes, anomalous floral organ formation, and large variations in the seed forming characteristics permitted us to determine the consequence of the extent of miR167 expression diminution accompanying the transcriptional biases of interrelated miRNAs. We demonstrate that percent diminution of miR167 gene expression is proportionally associated with both early and late flowering-time phenotypes in mimic lines. Also, the associated miRNAs, miR156, miR159, and miR172 showed >90% transcriptional diminution in at least ‘early-flowering’ miR167 mimic lines. On contrary, low percentages of their respective diminution were recorded in ‘late-flowering’ lines. Evidently, the misexpression of miR156, miR159, and miR172 led to the over-expression of their respective target genes SPL9, AtMYB33-like and AP2 genes in mimic lines which resulted in assorted phenotypes. We describe the scope of spatial regulation of these microRNAs in floral bud tissues of mimic lines which showed negative- or very low (<25%) misexpression levels in early/late-flowering lines highlighting their roles in the acquisition of flowering mechanism. To our knowledge, this study represents the first characterization of transcriptional biases of flowering associated miRNAs in miR167-mimic lines and certainly augments our understanding of the importance of microRNA-mediated regulation of flowering in plants.

Molecular pathways involved in microRNA-mediated regulation of multidrug resistance.

Multidrug resistance (MDR) is still a major reason for therapeutic failure in cancers. The mechanisms underlying MDR are multifactorial and are not fully understood. miRNAs are evolutionarily conserved non-coding RNAs that function as key post-transcriptional regulators of gene expression. Emerging evidence indicates a vital role of miRNAs in the resistance to cancer treatments and suggests their potential for cancer therapy, as they can modulate multiple mechanisms link to the development of MDR. This review summarizes the recent findings on the role of miRNAs in MDR, and highlights the molecular targets and mechanisms of miRNA regulated pathways in MDR. Finally, the challenges and prospects of miRNA-targeted therapies for reversing drug resistance are reviewed.

Pentafluoropropionic Anhydride Functionalized PAMAM Dendrimer as miRNA Delivery Reagent

Poly(amidoamine) (PAMAM) dendrimers are good candidates for nucleic acid delivery with their well-defined characteristics. MicroRNA (miRNA) mediated regulation of biological process are also active area of investigation. Fibroblast cells, such as MRC-5, are one of the cell lines used in biological researches due to their hard to transfect nature. In this two-staged study, cystamine core G5 PAMAM dendrimers were synthesized and modified with pentafluoropropionic anhydride (PA) and subsequently tested as miRNA delivery reagent on MRC-5 cells. Effect of fluorination against to naked G5 dendrimer over transfection efficiency was also investigated by molecular docking and quantitative structure-activity relationship (QSAR) calculations. Structural characterization of the synthesized dendrimers was verified by spectroscopic techniques. Gel retardation assay, particle size and transmission electron microscopy results demonstrated polyplex formation ability of fluorinated dendrimers with miRNA at nanoscale level. Zeta potential values indicated non-aggregation and increased stability of the polyplexes. Prepared polyplexes with fluorinated dendrimer showed over 90% cell viability and transfection efficiency. In silico calculations confirmed the stable complexation with miRNA and smooth penetration capability into the cell.

Genome-scale MicroRNA target prediction through clustering with Dirichlet process mixture model

BackgroundMicroRNA regulation is fundamentally responsible for fine-tuning the whole gene network in human and has been implicated in most physiological and pathological conditions. Studying regulatory impact of microRNA on various cellular and disease processes has resulted in numerous computational tools that investigate microRNA-mRNA interactions through the prediction of static binding site highly dependent on sequence pairing. However, what hindered the practical use of such target prediction is the interplay between competing and cooperative microRNA binding that complicates the whole regulatory process exceptionally.ResultsWe developed a new method for improved microRNA target prediction based on Dirichlet Process Gaussian Mixture Model (DPGMM) using a large collection of molecular features associated with microRNA, mRNA, and the interaction sites. Multiple validations based on microRNA-mRNA interactions reported in recent large-scale sequencing analyses and a screening test on the entire human transcriptome show that our model outperformed several state-of-the-art tools in terms of promising predictive power on binding sites specific to transcript isoforms with reduced false positive prediction. Last, we illustrated the use of predicted targets in constructing conditional microRNA-mediated gene regulation networks in human cancer.ConclusionThe probability-based binding site prediction provides not only a useful tool for differentiating microRNA targets according to the estimated binding potential but also a capability highly important for exploring dynamic regulation where binding competition is involved.