The virulome of Streptomyces scabiei in response to cello-oligosaccharide elicitors.

The development of spots or lesions symptomatic of common scab on root and tuber crops is caused by few pathogenic Streptomyces with Streptomyces scabiei 87-22 as the model species. Thaxtomin phytotoxins are the primary virulence determinants, mainly acting by impairing cellulose synthesis, and their production in S. scabiei is in turn boosted by cello-oligosaccharides released from host plants. In this work we aimed to determine which molecules and which biosynthetic gene clusters (BGCs) of the specialized metabolism of S. scabiei 87-22 show a production and/or a transcriptional response to cello-oligosaccharides. Comparative metabolomic analyses revealed that molecules of the virulome of S. scabiei induced by cellobiose and cellotriose include (i) thaxtomin and concanamycin phytotoxins, (ii) desferrioxamines, scabichelin and turgichelin siderophores in order to acquire iron essential for housekeeping functions, (iii) ectoine for protection against osmotic shock once inside the host, and (iv) bottromycin and concanamycin antimicrobials possibly to prevent other microorganisms from colonizing the same niche. Importantly, both cello-oligosaccharides reduced the production of the spore germination inhibitors germicidins thereby giving the 'green light' to escape dormancy and trigger the onset of the pathogenic lifestyle. For most metabolites - either with induced or reduced production - cellotriose was revealed to be a slightly stronger elicitor compared to cellobiose, supporting an earlier hypothesis which suggested the trisaccharide was the real trigger for virulence released from the plant cell wall through the action of thaxtomins. Interestingly, except for thaxtomins, none of these BGCs' expression seems to be under direct control of the cellulose utilization repressor CebR suggesting the existence of a yet unknown mechanism for switching on the virulome. Finally, a transcriptomic analysis revealed nine additional cryptic BGCs that have their expression awakened by cello-oligosaccharides, suggesting that other and yet to be discovered metabolites could be part of the virulome of S. scabiei.

Epromoters function as a hub to recruit key transcription factors required for the inflammatory response.

Gene expression is controlled by the involvement of gene-proximal (promoters) and distal (enhancers) regulatory elements. Our previous results demonstrated that a subset of gene promoters, termed Epromoters, work as bona fide enhancers and regulate distal gene expression. Here, we hypothesized that Epromoters play a key role in the coordination of rapid gene induction during the inflammatory response. Using a high-throughput reporter assay we explored the function of Epromoters in response to type I interferon. We find that clusters of IFNa-induced genes are frequently associated with Epromoters and that these regulatory elements preferentially recruit the STAT1/2 and IRF transcription factors and distally regulate the activation of interferon-response genes. Consistently, we identified and validated the involvement of Epromoter-containing clusters in the regulation of LPS-stimulated macrophages. Our findings suggest that Epromoters function as a local hub recruiting the key TFs required for coordinated regulation of gene clusters during the inflammatory response.

Transcriptomic Analyses Reveal 2 and 4 Family Members of Cytochromes P450 (CYP) Involved in LPS Inflammatory Response in Pharynx of Ciona robusta.

Cytochromes P450 (CYP) are enzymes responsible for the biotransformation of most endogenous and exogenous agents. The expression of each CYP is influenced by a unique combination of mechanisms and factors including genetic polymorphisms, induction by xenobiotics, and regulation by cytokines and hormones. In recent years, Ciona robusta, one of the closest living relatives of vertebrates, has become a model in various fields of biology, in particular for studying inflammatory response. Using an in vivo LPS exposure strategy, next-generation sequencing (NGS) and qRT-PCR combined with bioinformatics and in silico analyses, compared whole pharynx transcripts from naïve and LPS-exposed C. robusta, and we provide the first view of cytochrome genes expression and miRNA regulation in the inflammatory response induced by LPS in a hematopoietic organ. In C. robusta, cytochromes belonging to 2B,2C, 2J, 2U, 4B and 4F subfamilies were deregulated and miRNA network interactions suggest that different conserved and species-specific miRNAs are involved in post-transcriptional regulation of cytochrome genes and that there could be an interplay between specific miRNAs regulating both inflammation and cytochrome molecules in the inflammatory response in C. robusta.

Evolution of Polymyxin Resistance Regulates Colibactin Production in Escherichia coli.

The complex reservoir of metabolite-producing bacteria in the gastrointestinal tract contributes tremendously to human health and disease. Bacterial composition, and by extension gut metabolomic composition, is undoubtably influenced by the use of modern antibiotics. Herein, we demonstrate that polymyxin B, a last resort antibiotic, influences the production of the genotoxic metabolite colibactin from adherent-invasive Escherichia coli (AIEC) NC101. Colibactin can promote colorectal cancer through DNA double stranded breaks and interstrand cross-links. While the structure and biosynthesis of colibactin have been elucidated, chemical-induced regulation of its biosynthetic gene cluster and subsequent production of the genotoxin by E. coli are largely unexplored. Using a multiomic approach, we identified that polymyxin B stress enhances the abundance of colibactin biosynthesis proteins (Clb's) in multiple pks+ E. coli strains, including pro-carcinogenic AIEC, NC101; the probiotic strain, Nissle 1917; and the antibiotic testing strain, ATCC 25922. Expression analysis via qPCR revealed that increased transcription of clb genes likely contributes to elevated Clb protein levels in NC101. Enhanced production of Clb's by NC101 under polymyxin stress matched an increased production of the colibactin prodrug motif, a proxy for the mature genotoxic metabolite. Furthermore, E. coli with a heightened tolerance for polymyxin induced greater mammalian DNA damage, assessed by quantification of γH2AX staining in cultured intestinal epithelial cells. This study establishes a key link between the polymyxin B stress response and colibactin production in pks+ E. coli. Ultimately, our findings will inform future studies investigating colibactin regulation and the ability of seemingly innocuous commensal microbes to induce host disease.

Genome-wide characterization and expression analyses of the auxin/indole-3-acetic acid (Aux/IAA) gene family in barley (Hordeum vulgare L.).

Aux/IAA genes are early auxin-responsive genes and essential for auxin signaling transduction. There is little information about Aux/IAAs in the agriculturally important cereal, barley. Using in silico method, we identified and subsequently characterized 36 Aux/IAAs from the barley genome. Based on their genomic sequences and the phylogenic relationship with Arabidopsis and rice Aux/IAA, the 36 HvIAAs were categorized into two major groups and 14 subgroups. The indication of the presence or absence of these domains for the biological functions and acting mechanisms was discussed. The cis-element distributions in HvIAA promoters suggests that the HvIAAs expressions may not only regulated by auxin (the presence of AuxREs and TGA-element) but also by other hormones and developmental and environmental cues. We then studied the HvIAAs expression in response to NAA (1-Naphthaleneacetic acid) using quantitative real-time PCR (qRT-PCR). Like the promoter analysis, only 14 HvIAAs were upregulated by NAA over two-fold at 4 h. HvIAAs were clustered into three groups based on the spatiotemporal expression data. We confirmed by qRT-PCR that most HvIAAs, especially HvIAA3, HvIAA7, HvIAA8, HvIAA18, HvIAA24 and HvIAA34, are expressed in the developing barley spike compared within seedling, suggesting their roles in regulating spike development. Taken together, our data provide a foundation for further revealing the biological function of these HvIAAs.

Genome-wide identification and expression analysis of the NAC transcription factor family in tomato (Solanum lycopersicum) during aluminum stress.

BACKGROUND: The family of NAC proteins (NAM, ATAF1/2, and CUC2) represent a class of large plant-specific transcription factors. However, identification and functional surveys of NAC genes of tomato (Solanum lycopersicum) remain unstudied, despite the tomato genome being decoded for several years. This study aims to identify the NAC gene family and investigate their potential roles in responding to Al stress. RESULTS: Ninety-three NAC genes were identified and named in accordance with their chromosome location. Phylogenetic analysis found SlNACs are broadly distributed in 5 groups. Gene expression analysis showed that SlNACs had different expression levels in various tissues and at different fruit development stages. Cycloheximide treatment and qRT-PCR analysis indicated that SlNACs may aid regulation of tomato in response to Al stress, 19 of which were significantly up- or down-regulated in roots of tomato following Al stress. CONCLUSION: This work establishes a knowledge base for further studies on biological functions of SlNACs in tomato and will aid in improving agricultural traits of tomato in the future.

Patient-specific cancer genes contribute to recurrently perturbed pathways and establish therapeutic vulnerabilities in esophageal adenocarcinoma.

The identification of cancer-promoting genetic alterations is challenging particularly in highly unstable and heterogeneous cancers, such as esophageal adenocarcinoma (EAC). Here we describe a machine learning algorithm to identify cancer genes in individual patients considering all types of damaging alterations simultaneously. Analysing 261 EACs from the OCCAMS Consortium, we discover helper genes that, alongside well-known drivers, promote cancer. We confirm the robustness of our approach in 107 additional EACs. Unlike recurrent alterations of known drivers, these cancer helper genes are rare or patient-specific. However, they converge towards perturbations of well-known cancer processes. Recurrence of the same process perturbations, rather than individual genes, divides EACs into six clusters differing in their molecular and clinical features. Experimentally mimicking the alterations of predicted helper genes in cancer and pre-cancer cells validates their contribution to disease progression, while reverting their alterations reveals EAC acquired dependencies that can be exploited in therapy.

Comparative analysis of B-BOX genes and their expression pattern analysis under various treatments in Dendrobium officinale.

BACKGROUND: Studies have demonstrated that BBX (B-BOX) genes play crucial roles in regulatory networks controlling plant growth, developmental processes and stress response. Nevertheless, comprehensive study of BBX genes in orchids (Orchidaceae) is not well studied. The newly released genome sequences of Dendrobium officinale and Phalaenopsis equestris have allowed a systematic analysis of these important BBX genes in orchids. RESULTS: Here we identified 19 (DoBBX01-19) and 16 (PeBBX01-16) BBX genes from D. officinale and P. equestris, respectively, and clustered into five clades (I-V) according to phylogenetic analysis. Thirteen orthologous, two DoBBXs paralogous and two PeBBXs paralogous gene pairs were validated. This gene family mainly underwent purifying selection, but five domains experienced positive selection during evolution. Noteworthy, the expression patterns of root, root_tips, stem, leaf, speal, column, lip, and flower_buds revealed that they might contribution to the formation of these tissues. According to the cis-regulatory elements analysis of BBX genes, qRT-PCR experiments were carried out using D. officinale PLBs (protocorm-like bodies) and displayed that these BBX genes were differentially regulated under AgNO3, MeJA (Methyl Jasmonate), ABA (abscisic acid) and SA (salicylic acid) treatments. CONCLUSIONS: Our analysis exposed that DoBBX genes play significant roles in plant growth and development, and response to different environmental stress conditions of D. officinale, which provide aid in the selection of appropriate candidate genes for further functional characterization of BBX genes in plants.

Mycobacterium tuberculosis cysteine biosynthesis genes mec+-cysO-cysM confer resistance to clofazimine.

The Mycobacterium tuberculosis mec+-cysO-cysM gene cluster was shown to be part of a novel cysteine biosynthesis pathway in vitro, but little is known about its essentiality or role in M. tuberculosis physiology. In this study, we generate a knock out of the mec+-cysO-cysM gene cluster in M. tuberculosis and show that the gene cluster is not essential under a variety of conditions, suggesting redundancy in pathways for cysteine biosynthesis in M. tuberculosis. The cysteine biosynthesis gene cluster is essential for resistance for clofazimine, a peroxide-producing anti-leprosy drug. Therefore, although under most conditions the pathway is not essential, it likely has an important role in defense against oxidative stress in M. tuberculosis.

The effects of valproic acid on early pregnancy human placentas: Pilot ex vivo analysis in cultured placental villi.

Valproic acid is an established structural and neurodevelopmental teratogen. Recently, we demonstrated that valproate alters the barrier function of perfused term human placentas. Here, we conducted a pilot study to evaluate the effects of subchronic valproate exposure on carrier expression in cultured placental villous explants from early human pregnancies. Placental tissue of gestational age 6-13 weeks was collected from elective pregnancy terminations in women without known epilepsy. The effects of valproate (42, 83, or 166 µg/mL) on the mRNA expression of 37 major placental carriers and related genes were evaluated by a customized gene expression array (n = 5, 5 days). Five-day exposure to valproate was associated with high variability in gene expression. However, two main gene clusters were identified, including a cluster of three major folate carriers. Exposure to low therapeutic levels of valproate (42 µg/mL) was associated with a tendency toward reduced mRNA expression of genes encoding folate and amino acid and fatty acid carriers (P = 0.065, paired analysis). Our initial findings suggest that valproate can affect the function of the human placenta during early pregnancy.

Sulfur dioxide response of Brettanomyces bruxellensis strains isolated from Greek wine.

Brettanomyces bruxellensis is the most common spoilage wine yeast which can provoke great economic damage to the wine industry due to the production of undesirable odors. The capacity of the species to adapt in various environmental conditions offers a selective advantage that is reflected by intraspecific variability at genotypic and phenotypic level. In this study, microsatellite analysis of 22 strains isolated from Greek wine revealed the existence of distinct genetic subgroups that are correlated with their geographical origin. The response of these strains to increasing levels of sulfur dioxide confirmed the presence of both sensitive and tolerant strains, which belong to distinguished genetic clusters. The genetic categorization of B. bruxellensis strains could be used by the winemakers as a diagnostic tool regarding sulfur dioxide sensitivity.

Targeting EIF4E signaling with ribavirin in infant acute lymphoblastic leukemia.

The poor outcomes in infant acute lymphoblastic leukemia (ALL) necessitate new treatments. Here we discover that EIF4E protein is elevated in most cases of infant ALL and test EIF4E targeting by the repurposed antiviral agent ribavirin, which has anticancer properties through EIF4E inhibition, as a potential treatment. We find that ribavirin treatment of actively dividing infant ALL cells on bone marrow stromal cells (BMSCs) at clinically achievable concentrations causes robust proliferation inhibition in proportion with EIF4E expression. Further, we find that ribavirin treatment of KMT2A-rearranged (KMT2A-R) infant ALL cells and the KMT2A-AFF1 cell line RS4:11 inhibits EIF4E, leading to decreases in oncogenic EIF4E-regulated cell growth and survival proteins. In ribavirin-sensitive KMT2A-R infant ALL cells and RS4:11 cells, EIF4E-regulated proteins with reduced levels of expression following ribavirin treatment include MYC, MCL1, NBN, BCL2 and BIRC5. Ribavirin-treated RS4:11 cells exhibit impaired EIF4E-dependent nuclear to cytoplasmic export and/or translation of the corresponding mRNAs, as well as reduced phosphorylation of the p-AKT1, p-EIF4EBP1, p-RPS6 and p-EIF4E signaling proteins. This leads to an S-phase cell cycle arrest in RS4:11 cells corresponding to the decreased proliferation. Ribavirin causes nuclear EIF4E to re-localize to the cytoplasm in KMT2A-AFF1 infant ALL and RS4:11 cells, providing further evidence for EIF4E inhibition. Ribavirin slows increases in peripheral blasts in KMT2A-R infant ALL xenograft-bearing mice. Ribavirin cooperates with chemotherapy, particularly L-asparaginase, in reducing live KMT2A-AFF1 infant ALL cells in BMSC co-cultures. This work establishes that EIF4E is broadly elevated across infant ALL and that clinically relevant ribavirin exposures have preclinical activity and effectively inhibit EIF4E in KMT2A-R cases, suggesting promise in EIF4E targeting using ribavirin as a means of treatment.

miR-17-92 cluster is connected with disease progression and oxaliplatin/capecitabine chemotherapy efficacy in advanced gastric cancer patients: A preliminary study.

This study aimed to determine the role of plasma miR-17-92 cluster level in predicting chemoresistance in patients with gastric cancer (GC) undergoing oxaliplatin/capecitabine (XELOX) chemotherapy.Patients recently diagnosed with advanced GC were chosen as participants based on the inclusion criteria. The plasma levels of miR-17-5p, miR-18a, miR-19a/b, miR-20a, and miR-92-1 (miR-17-92 cluster) were determined through quantitative RT-PCR of blood samples from GC patients and healthy volunteers. All the patients received XELOX chemotherapy, and the effectiveness of the chemotherapy was evaluated.The miR-17-92 plasma level was increased in advanced GC patients and decreased after XELOX chemotherapy. Moreover, the miR-17-92 cluster level was associated with chemotherapy response but not with chemotherapy-related toxicity. The miR-17-92 cluster plasma level was decreased in chemosensitive patients, but not in chemoresistant patients, after chemotherapy. The sensitivity and specificity of the combined detection of the miR-17-92 cluster in patients with advanced GC were 100% each.The results suggest that the miR-17-92 plasma level is associated with the progression of advanced GC and effectiveness of XELOX chemotherapy.

Cracking the regulatory code of biosynthetic gene clusters as a strategy for natural product discovery.

The World Health Organization (WHO) describes antibiotic resistance as "one of the biggest threats to global health, food security, and development today", as the number of multi- and pan-resistant bacteria is rising dangerously. Acquired resistance phenomena also impair antifungals, antivirals, anti-cancer drug therapy, while herbicide resistance in weeds threatens the crop industry. On the positive side, it is likely that the chemical space of natural products goes far beyond what has currently been discovered. This idea is fueled by genome sequencing of microorganisms which unveiled numerous so-called cryptic biosynthetic gene clusters (BGCs), many of which are transcriptionally silent under laboratory culture conditions, and by the fact that most bacteria cannot yet be cultivated in the laboratory. However, brute force antibiotic discovery does not yield the same results as it did in the past, and researchers have had to develop creative strategies in order to unravel the hidden potential of microorganisms such as Streptomyces and other antibiotic-producing microorganisms. Identifying the cis elements and their corresponding transcription factors(s) involved in the control of BGCs through bioinformatic approaches is a promising strategy. Theoretically, we are a few 'clicks' away from unveiling the culturing conditions or genetic changes needed to activate the production of cryptic metabolites or increase the production yield of known compounds to make them economically viable. In this opinion article, we describe and illustrate the idea beyond 'cracking' the regulatory code for natural product discovery, by presenting a series of proofs of concept, and discuss what still should be achieved to increase the rate of success of this strategy.

Piperine inhibits aflatoxin B1 production in Aspergillus flavus by modulating fungal oxidative stress response.

Aspergillus flavus, a soil-borne pathogen, represents a danger for humans and animals since it produces the carcinogenic mycotoxin Aflatoxin B1 (AFB1). Approaches aiming the reduction of this fungal contaminant mainly involve chemicals that may also be toxic. Therefore, identification and characterization of natural anti-aflatoxigenic products represents a sustainable alternative strategy. Piperine, a major component of black and long peppers, has been previously demonstrated asan AFB1-inhibitor; nevertheless its mechanism of action was yet to be elucidated. The aim of the present study was to evaluate piperine's molecular mechanism of action in A. flavus with a special focus on oxidative stress response. For that, the entire AFB1 gene cluster as well asa targeted gene-network coding for fungal stress response factors and cellular receptors were analyzed. In addition to this, fungal enzymatic activities were also characterized. We demonstrated that piperine inhibits aflatoxin production and fungal growth in a dose-dependent manner. Analysis of the gene cluster demonstrated that almost all genes participating in aflatoxin's biosynthetic pathway were down regulated. Exposure to piperine also resulted in decreased transcript levels of the global regulator veA together with an over-expression of genes coding for several basic leucine zipper (bZIP) transcription factors such as atfA, atfB and ap-1 and genes belonging to superoxide dismutase and catalase's families. Furthermore, this gene response was accompanied by a significant enhancement of catalase enzymatic activity. In conclusion, these data demonstrated that piperine inhibits AFB1 production while positively modulating fungal antioxidant status in A. flavus.

At the base of colinear Hox gene expression: cis-features and trans-factors orchestrating the initial phase of Hox cluster activation.

Hox genes are crucial players in the generation and pattering of the vertebrate trunk and posterior body during embryogenesis. Their initial expression takes place shortly after the establishment of the primitive streak, in the posterior-most part of the mouse embryo and is a determinant step for setting up the definitive Hox expression boundaries along the antero-posterior body axis. The developmental signals and epigenetic mechanisms underlying this early activation remained unsolved until recently. The development of novel embryo-derived model systems, combined with methods that examine chromatin status and chromosome conformation, led to deeper understanding of the process of Hox activation in the early embryo. Here we summarize how the early Hox cis-regulatory landscape becomes active upon receiving the appropriate developmental signal, and we discuss the importance of the local topological segmentation of the HoxA cluster during early Hox activation.

Genome-wide analysis and expression characteristics of small auxin-up RNA (SAUR) genes in moso bamboo (Phyllostachys edulis).

Moso bamboo (Phyllostachys edulis) is well known for its rapid shoot growth. Auxin exerts pleiotropic effects on plant growth. The small auxin-up RNA (SAUR) genes are early auxin-responsive genes involved in plant growth. In total, 38 SAUR genes were identified in P. edulis (PheSAUR). A comprehensive overview of the PheSAUR gene family is presented, including the gene structures, phylogeny, and subcellular location predictions. A transcriptome analysis indicated that 37 (except PheSAUR18) of the PheSAUR genes were expressed during shoot growth process and that the PheSAUR genes were differentially expressed. Furthermore, quantitative real-time PCR analysis indicated that all of the PheSAUR genes could be induced in different tissues of seedlings and that 37 (except PheSAUR41) of the PheSAUR genes were up-regulated after indole-3-acetic acid (IAA) treatment. These results reveal a comprehensive overview of the PheSAUR gene family and may pave the way for deciphering their functions during bamboo development.

Transcriptome Profile Reveals that Pu-Erh Tea Represses the Expression of Vitellogenin Family to Reduce Fat Accumulation in Caenorhabditis elegans.

Due to misbalanced energy surplus and expenditure, obesity has become a common chronic disorder that is highly associated with many metabolic diseases. Pu-erh tea, a traditional Chinese beverage, has been believed to have numerous health benefits, such as anti-obesity. However, the underlying mechanisms of its anti-obesity effect are yet to be understood. Here, we take the advantages of transcriptional profile by RNA sequencing (RNA-Seq) to view the global gene expression of Pu-erh tea. The model organism Caenorhabditis elegans was treated with different concentrations of Pu-erh tea water extract (PTE, 0 g/mL, 0.025 g/mL, and 0.05 g/mL). Compared with the control, PTE indeed decreases lipid droplets size and fat accumulation. The high-throughput RNA-Sequence technique detected 18073 and 18105 genes expressed in 0.025 g/mL and 0.05 g/mL PTE treated groups, respectively. Interestingly, the expression of the vitellogenin family (vit-1, vit-2, vit-3, vit-4 and vit-5) was significantly decreased by PTE, which was validated by qPCR analysis. Furthermore, vit-1(ok2616), vit-3(ok2348) and vit-5(ok3239) mutants are insensitive to PTE triggered fat reduction. In conclusion, our transcriptional profile by RNA-Sequence suggests that Pu-erh tea lowers the fat accumulation primarily through repression of the expression of vit(vitellogenin) family, in addition to our previously reported (sterol regulatory element binding protein) SREBP-SCD (stearoyl-CoA desaturase) axis.

Design of a miRNA sponge for the miR-17 miRNA family as a therapeutic strategy against vulvar carcinoma.

Dysregulation of microRNAs has been studied thoroughly, and has been observed in a variety of tumors including vulvar carcinomas, a rare type of gynecological tumor with increasing incidence. However, very few therapeutic alternatives have reached the clinical setting, and there is an urgent unmet need to develop novel strategies for patients with this tumor type. Thus, a microRNA (miRNA) sponge for the miR-17 miRNA family was designed, synthesized and validated in vitro in order to explore a new therapeutic strategy based on inhibiting this oncogenic miRNA family in vulvar cancer. Members of the miR-17 family were evaluated for expression in a vulvar tumor cell line (SW954) and 20 HPV negative formalin-fixed paraffin-embedded (FFPE) samples by quantitative real-time PCR (qRT-PCR). Six in tandem, bulged sequences that were complementary to these miRNAs were designed, synthesized, cloned, and transfected into SW954 cells. A luciferase reporter assay with a psiCheck2 vector was used to test the specificity of the sponge sequences for miR-17 family miRNA binding. Taqman qRT-PCR was used to test how the sponges affected miRNA expression. In FFPE samples, higher expression of miR-20a and miR-106a correlated with deeper tumor invasion (P = 0.0187 and P = 0.0404, respectively). The luciferase reporter assay validated the specificity of the sponge for miR-17 family members. Using qRT-PCR, we confirmed this specificity with decreased expression in 5 (out of six) miRNAs of the miR-17 family in SW954 cells. Although our results are preliminary, these results demonstrate that these miRNA sponges are potent inhibitors of the miR-17 family of miRNAs in SW954. Therefore, this miRNA-specific sponge may be developed into a novel therapeutic treatment for patients with vulvar cancer.

Integrative Analysis of MicroRNA and mRNA Data Reveals an Orchestrated Function of MicroRNAs in Skeletal Myocyte Differentiation in Response to TNF-α or IGF1.

INTRODUCTION: Skeletal muscle cell differentiation is impaired by elevated levels of the inflammatory cytokine tumor necrosis factor-α (TNF-α) with pathological significance in chronic diseases or inherited muscle disorders. Insulin like growth factor-1 (IGF1) positively regulates muscle cell differentiation. Both, TNF-α and IGF1 affect gene and microRNA (miRNA) expression in this process. However, computational prediction of miRNA-mRNA relations is challenged by false positives and targets which might be irrelevant in the respective cellular transcriptome context. Thus, this study is focused on functional information about miRNA affected target transcripts by integrating miRNA and mRNA expression profiling data. METHODOLOGY/PRINCIPAL FINDINGS: Murine skeletal myocytes PMI28 were differentiated for 24 hours with concomitant TNF-α or IGF1 treatment. Both, mRNA and miRNA expression profiling was performed. The data-driven integration of target prediction and paired mRNA/miRNA expression profiling data revealed that i) the quantity of predicted miRNA-mRNA relations was reduced, ii) miRNA targets with a function in cell cycle and axon guidance were enriched, iii) differential regulation of anti-differentiation miR-155-5p and miR-29b-3p as well as pro-differentiation miR-335-3p, miR-335-5p, miR-322-3p, and miR-322-5p seemed to be of primary importance during skeletal myoblast differentiation compared to the other miRNAs, iv) the abundance of targets and affected biological processes was miRNA specific, and v) subsets of miRNAs may collectively regulate gene expression. CONCLUSIONS: Joint analysis of mRNA and miRNA profiling data increased the process-specificity and quality of predicted relations by statistically selecting miRNA-target interactions. Moreover, this study revealed miRNA-specific predominant biological implications in skeletal muscle cell differentiation and in response to TNF-α or IGF1 treatment. Furthermore, myoblast differentiation-associated miRNAs are suggested to collectively regulate gene clusters and targets associated with enriched specific gene ontology terms or pathways. Predicted miRNA functions of this study provide novel insights into defective regulation at the transcriptomic level during myocyte proliferation and differentiation due to inflammatory stimuli.