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Abstract Therapy of melanoma has improved dramatically over the last years thanks to the development of targeted therapies (MAPKi) and immunotherapies. However, drug resistance continues to limit the efficacy of these therapies. Our research group has provided robust evidence as to the involvement of a set of microRNAs in the development of resistance to target therapy in BRAF-mutated melanomas. Among them, a pivotal role is played by the oncosuppressor miR-579-3p. Here we show that miR-579-3p and the microphthalmia-associated transcription factor (MITF) influence reciprocally their expression through positive feedback regulatory loops. In particular we show that miR-579-3p is specifically deregulated in BRAF-mutant melanomas and that its expression levels mirror those of MITF. Luciferase and ChIP studies show that MITF is a positive regulator of miR-579-3p, which is located in the intron 11 of the human gene ZFR (Zink-finger recombinase) and is co-transcribed with its host gene. Moreover, miR-579-3p, by targeting BRAF, is able to stabilize MITF protein thus inducing its own transcription. From biological points of view, early exposure to MAPKi or, alternatively miR-579-3p transfection, induce block of proliferation and trigger senescence programs in BRAF-mutant melanoma cells. Finally, the long-term development of resistance to MAPKi is able to select cells characterized by the loss of both miR-579-3p and MITF and the same down-regulation is also present in patients relapsing after treatments. Altogether these findings suggest that miR-579-3p/MITF interplay potentially governs the balance between proliferation, senescence and resistance to therapies in BRAF-mutant melanomas.

The focus of this paper is to evaluate how well European countries are advancing towards a Circular Economy transition and to which extent they are fulfilling the European Union (EU) requirements in terms of municipal waste management. With this aim, an innovative composite indicator is devised by integrating the Goal Programming Synthetic Indicator (GPSI) methodology with the Analytic Hierarchy Process (AHP). The suggested methodology allows to encompass not only the guidelines provided by the EU directives and the EU Waste Hierarchy, but also the deviations from the EU thresholds. As a first step of the analysis, a dashboard of indicators is identified together with a set of targets that European countries are supposed to meet. Indicators, targets and their relative importance in the dashboard have been discussed and validated by a team of experts. Then, two Goal Programming Synthetic Indicators are computed taking into account two different perspectives. The first one rewards the countries with good performance in the higher level of the Waste Hierarchy, while the second one penalizes countries whose infringements are in the lower part of the Waste Hierarchy. Hence two different systems of aggregating weights are identified by means of the Analytic Hierarchy Process and accordingly two scenarios are explored. The analysis is performed using Eurostat data on 28 European countries from 2013 to 2018. For each year, countries are assessed in terms of their ability to keep the right waste management track delineated by the Waste Hierarchy principles. Countries’ ranking over time is first obtained and then interpreted in light of countries policies and achievements, deriving policy suggestions to improve waste management strategy able to reach the expected results.

Abstract Aging progressively modifies the physiological balance of the organism increasing susceptibility to both genetic and sporadic neurodegenerative diseases. These changes include epigenetic chromatin remodeling events that may modify the transcription levels of disease-causing genes affecting neuronal survival. However, how these events interconnect is not well understood. Here, we found that Su(var)3-9 causes increased methylation of histone H3K9 in the promoter region of TDP-43, the most frequently altered factor in amyotrophic lateral sclerosis (ALS), affecting the mRNA and protein expression levels of this gene through epigenetic modifications that appear to be conserved in aged Drosophila brains, mouse, and human cells. Remarkably, augmented Su(var)3-9 activity causes a decrease in TDP-43 expression followed by early defects in locomotor activities. In contrast, decreasing Su(var)3-9 action promotes higher levels of TDP-43 expression, improving motility parameters in old flies. The data uncover a novel role of this enzyme in regulating TDP-43 expression and locomotor senescence and indicate conserved epigenetic mechanisms that may play a role in the pathogenesis of ALS.

Familial Dysautonomia (FD) is a rare disease caused by ELP1 exon 20 skipping. Here we clarify the role of RNA Polymerase II (RNAPII) and chromatin on this splicing event. A slow RNAPII mutant and chromatin-modifying chemicals that reduce the rate of RNAPII elongation induce exon skipping whereas chemicals that create a more relaxed chromatin exon inclusion. In the brain of a mouse transgenic for the human FD-ELP1 we observed on this gene an age-dependent decrease in the RNAPII density profile that was most pronounced on the alternative exon, a robust increase in the repressive marks H3K27me3 and H3K9me3 and a decrease of H3K27Ac, together with a progressive reduction in ELP1 exon 20 inclusion level. In HEK 293T cells, selective drug-induced demethylation of H3K27 increased RNAPII elongation on ELP1 and SMN2, promoted the inclusion of the corresponding alternative exons, and, by RNA-sequencing analysis, induced changes in several alternative splicing events. These data suggest a co-transcriptional model of splicing regulation in which age-dependent changes in H3K27me3/Ac modify the rate of RNAPII elongation and affect processing of ELP1 alternative exon 20.

IntroductionSmall cell lung cancer (SCLC) is characterized by poor prognosis and challenging diagnosis. Screening in high-risk smokers results in a reduction in lung cancer mortality, however, screening efforts are primarily focused on non-small cell lung cancer (NSCLC). SCLC diagnosis and surveillance remain significant challenges. The aberrant expression of circulating microRNAs (miRNAs/miRs) is reported in many tumors and can provide insights into the pathogenesis of tumor development and progression. Here, we conducted a comprehensive assessment of circulating miRNAs in SCLC with a goal of developing a miRNA-based classifier to assist in SCLC diagnoses.MethodsWe profiled deregulated circulating cell-free miRNAs in the plasma of SCLC patients. We tested selected miRNAs on a training cohort and created a classifier by integrating miRNA expression and patients’ clinical data. Finally, we applied the classifier on a validation dataset.ResultsWe determined that miR-375-3p can discriminate between SCLC and NSCLC patients, and between SCLC and Squamous Cell Carcinoma patients. Moreover, we found that a model comprising miR-375-3p, miR-320b, and miR-144-3p can be integrated with race and age to distinguish metastatic SCLC from a control group.DiscussionThis study proposes a miRNA-based biomarker classifier for SCLC that considers clinical demographics with specific cut offs to inform SCLC diagnosis.

Abstract Alterations in the function of the RNA-binding protein TDP-43 are largely associated with the pathogenesis of amyotrophic lateral sclerosis (ALS), a devastating disease of the human motor system that leads to motoneurons degeneration and reduced life expectancy by molecular mechanisms not well known. In our previous work, we found that the expression levels of the glutamic acid decarboxylase enzyme (GAD1), responsible for converting glutamate to γ-aminobutyric acid (GABA), were downregulated in TBPH-null flies and motoneurons derived from ALS patients carrying mutations in TDP-43, suggesting that defects in the regulation of GAD1 may lead to neurodegeneration by affecting neurotransmitter balance. In this study, we observed that TBPH was required for the regulation of GAD1 pre-mRNA splicing and the levels of GABA in the Drosophila central nervous system (CNS). Interestingly, we discovered that pharmacological treatments aimed to potentiate GABA neurotransmission were able to revert locomotion deficiencies in TBPH-minus flies, revealing novel mechanisms and therapeutic strategies in ALS.

Abstract MicroRNAs (miRNAs) are regulatory small non-coding RNAs that function as translational repressors. MiRNAs are involved in most cellular processes, and their expression and function are presided by several factors. Amongst, miRNA editing is an epitranscriptional modification that alters the original nucleotide sequence of selected miRNAs, possibly influencing their biogenesis and target-binding ability. A-to-I and C-to-U RNA editing are recognized as the canonical types, with the A-to-I type being the predominant one. Albeit some bioinformatics resources have been implemented to collect RNA editing data, it still lacks a comprehensive resource explicitly dedicated to miRNA editing. Here, we present MiREDiBase, a manually curated catalog of editing events in miRNAs. The current version includes 3,059 unique validated and putative editing sites from 626 pre-miRNAs in humans and three primates. Editing events in mature human miRNAs are supplied with miRNA-target predictions and enrichment analysis, while minimum free energy structures are inferred for edited pre-miRNAs. MiREDiBase represents a valuable tool for cell biology and biomedical research and will be continuously updated and expanded at https://ncrnaome.osumc.edu/miredibase .

miRNAs are some of the most well-characterized regulators of gene expression. Integral to several physiological processes, their aberrant expression often drives the pathogenesis of both benign and malignant diseases. Similarly, DNA methylation represents an epigenetic modification influencing transcription and playing a critical role in silencing numerous genes. The silencing of tumor suppressor genes through DNA methylation has been reported in many types of cancer and is associated with tumor development and progression. A growing body of literature has described the crosstalk between DNA methylation and miRNAs as an additional layer in the regulation of gene expression. Methylation in miRNA promoter regions inhibits its transcription, while miRNAs can target transcripts and subsequently regulate the proteins responsible for DNA methylation. Such relationships between miRNA and DNA methylation serve an important regulatory role in several tumor types and highlight a novel avenue for potential therapeutic targets. In this review, we discuss the crosstalk between DNA methylation and miRNA expression in the pathogenesis of cancer and describe how miRNAs influence DNA methylation and, conversely, how methylation impacts the expression of miRNAs. Finally, we address how these epigenetic modifications may be leveraged as biomarkers in cancer.

Abstract Background Mutations in the small RNA-binding protein TDP-43 lead to the formation of insoluble cytoplasmic aggregates that have been associated with the onset and progression of amyotrophic lateral sclerosis (ALS), a neurodegenerative disorder affecting homeostasis of the motor system which is also characterized by aberrant expression of retrotransposable elements (RTEs). Although the TDP-43 function was shown to be required in the neurons and glia to maintain the organization of neuromuscular synapses and prevent denervation of the skeletal muscles, the molecular mechanisms involved in physiological dysregulation remain elusive. Here, we address this issue using a null mutation of the TDP-43 Drosophila homolog, TBPH. Results Using genome-wide gene expression profiles, we detected a strong upregulation of RTE expression in TBPH-null Drosophila heads, while the genetic rescue of the TDP-43 function reverted these modifications. Furthermore, we found that TBPH modulates the small interfering RNA (siRNA) silencing machinery responsible for RTE repression. Molecularly, we observed that TBPH regulates the expression levels of Dicer-2 by direct protein-mRNA interactions in vivo. Accordingly, the genetic or pharmacological recovery of Dicer-2 activity was sufficient to repress retrotransposon activation and promote motoneuron axonal wrapping and synaptic growth in TBPH-null Drosophila. Conclusions We identified an upregulation of RTE expression in TBPH-null Drosophila heads and demonstrate that defects in the siRNA pathway lead to RTE upregulation and motoneuron degeneration. Our results describe a novel physiological role of endogenous TDP-43 in the prevention of RTE-induced neurological alterations through the modulation of Dicer-2 activity and the siRNA pathway.

Abstract Background The ribonuclear protein TDP-43 has been implicated in the pathophysiology of amyotrophic lateral sclerosis (ALS), with genetic mutations being linked to the neurological symptoms of the disease. Though alterations in the intracellular distribution of TDP-43 have been observed in skeletal muscles of patients suffering from ALS, it is not clear whether such modifications play an active role in the disease or merely represent an expression of muscle homeostatic mechanisms. Also, the molecular and metabolic pathways regulated by TDP-43 in the skeletal muscle remain largely unknown. Here, we analyze the function of TBPH, the Drosophila melanogaster ortholog of TDP-43, in skeletal muscles. Results We modulated the activity of TDP-43 in Drosophila muscles by means of RNA interference and observed that it is required to promote the formation and growth of neuromuscular synapses. TDP-43 regulated the expression levels of Disc-large (Dlg), and restoring Dlg expression either in skeletal muscles or in motoneurons was sufficient to suppress the locomotive and synaptic defects of TDP-43-null flies. These results were validated by the observation of a decrease in Dlg levels in human neuroblastoma cells and iPSC-differentiated motoneurons derived from ALS patients, suggesting similar mechanisms may potentially be involved in the pathophysiology of the disease. Conclusions Our results help to unveil the physiological role of TDP-43 in skeletal muscles as well as the mechanisms responsible for the autonomous and non-autonomous behavior of this protein concerning the organization of neuromuscular synapses.

To determine the molecular and functional interactions between RNA-binding proteins (RBPs) and their targets RNAs, is of fundamental importance to understand the dynamic organization of the nervous system in health and disease. Nevertheless, this task has remained elusive due to the lack of specific protocols and experimental systems that would allow the combination of biochemical analysis with in vivo functional genetics. In this manuscript, we describe a trustworthy and detailed methodology to establish the molecular organization and intracellular function of RBPs/RNA multimeric complexes in a cell type-defined manner by using the powerful GAL4/UAS system for gene expression in Drosophila melanogaster.Graphic abstract: Immunoprecipitation for protein-RNA interaction in Drosophila.

Lung and breast cancer are the two most common causes of malignant pleural effusion (MPE). MPE diagnosis plays a crucial role in determining staging and therapeutic interventions in these cancers. However, our understanding of the pathogenesis and progression of MPE at the molecular level is limited. Extracellular Vesicles (EVs) and their contents, including microRNAs (miRNAs), can be isolated from all bodily fluids, including pleural fluid. This study aims to compare EV-miRNA patterns of expression in MPE caused by breast (BA-MPE) and lung (LA-MPE) adenocarcinomas compared to the control group of heart-failure-induced effusions (HF-PE). We conducted an analysis of 24 pleural fluid samples (8 LA-MPE, 8 BA-MPE, and 8 HF-PE). Using NanoString technology, we profiled miRNAs within EVs isolated from 12 cases. Bioinformatic analysis demonstrated differential expression of miR-1246 in the MPE group vs. HF-PE group and miR-150-5p and miR-1246 in the BA-MPE vs. LA-MPE group, respectively. This difference was demonstrated and validated in an independent cohort using real-time PCR (RT-PCR). miRNA-1246 demonstrated 4-fold increased expression (OR: 3.87, 95% CI: 0.43, 35) in the MPE vs. HF-PE group, resulting in an area under the curve of 0.80 (95% CI: 0.60, 0.99). The highest accuracy for differentiating MPE vs. HF-PE was seen with a combination of miRNAs compared to each miRNA alone. Consistent with prior studies, this study demonstrates dysregulation of specific EV-based miRNAs in breast and lung cancer; pleural fluid provides direct access for the analysis of these EV-miRNAs as biomarkers and potential targets and may provide insight into the underlying pathogenesis of tumor progression. These findings should be explored in large prospective studies.

Objective: to unveil the perceptions of the Street Clinic nursing staff about coping with vulnerabilities. Methods: qualitative study, carried out by means of participant observation of the team’s activities, recording in a field diary and semi-structured interviews with the nursing team, totaling 17 participants. Results: situations experienced by people living on the streets that deepen health inequities by violating rights were revealed. Among the nursing work tools, the potential of collaborative work, listening, and welcoming technologies stand out as mediators of a more humanized care. There is a need for specific strategies to guide nursing care on the streets. Conclusion: nursing has great potential for addressing the vulnerabilities of the homeless population using soft and soft-hard technologies.

The epitranscriptome encompasses all post-transcriptional modifications that occur on RNAs. These modifications can alter the function and regulation of their RNA targets, which, if dysregulated, result in various diseases and cancers. As with other RNAs, miRNAs are highly modified by epitranscriptomic modifications such as m6A methylation, 2′-O-methylation, m5C methylation, m7G methylation, polyuridine, and A-to-I editing. miRNAs are a class of small non-coding RNAs that regulates gene expression at the post-transcriptional level. miRNAs have gathered high clinical interest due to their role in disease, development, and cancer progression. Epitranscriptomic modifications alter the targeting, regulation, and biogenesis of miRNAs, increasing the complexity of miRNA regulation. In addition, emerging studies have revealed crosstalk between these modifications. In this review, we will summarize the epitranscriptomic modifications—focusing on those relevant to miRNAs—examine the recent crosstalk between these modifications, and give a perspective on how this crosstalk expands the complexity of miRNA biology.

Alterations in synaptic transmission are critical early events in neuromuscular disorders. However, reliable methodologies to analyze the functional organization of the neuromuscular synapses are still needed. This manuscript provides a detailed protocol to analyze the molecular assembly of the neuromuscular synapses through immune-electrophysiology in Drosophila melanogaster. This technique allows the quantification of the molecular behavior of the neuromuscular synapses by correlating the structural configuration of the synaptic boutons with their electrical activity.

Despite the development of targeted therapeutics, immunotherapy, and strategies for early detection, lung cancer carries a high mortality. Further, significant racial disparities in outcomes exist for which the molecular drivers have yet to be fully elucidated. The growing field of Epitranscriptomics has introduced a new layer of complexity to the molecular pathogenesis of cancer. RNA modifications can occur in coding and non-coding RNAs, such as miRNAs, possibly altering their gene regulatory function. The potential role for such modifications as clinically informative biomarkers remains largely unknown. Here, we concurrently profiled canonical miRNAs, shifted isomiRs (templated and non-templated), and miRNAs with single-point modification events (RNA and DNA) in White American (W) and Black or African American (B/AA) lung adenocarcinoma (LUAD) patients. We found that while most deregulated miRNA isoforms were similar in W and B/AA LUAD tissues compared to normal adjacent tissues, there was a subgroup of isoforms with deregulation according to race. We specifically investigated an edited miRNA, miR-151a-3p with an A-to-I editing event at position 3, to determine how its altered expression may be associated with activation of divergent biological pathways between W and B/AA LUAD patients. Finally, we identified distinct race-specific miRNA isoforms that correlated with prognosis for both Ws and B/AAs. Our results suggested that concurrently profiling canonical and non-canonical miRNAs may have potential as a strategy for identifying additional distinct biological pathways and biomarkers in lung cancer.

Lung cancer is the leading cause of cancer mortality worldwide. Increased understanding of the molecular mechanisms of the disease has led to the development of novel therapies and improving outcomes. Recently, extracellular vesicles (EVs) have emerged as vehicles for the transfer of genetic information between tumors and their microenvironment and have been implicated in lung cancer initiation, progression, and response to therapy. However, the mechanisms that drive the biogenesis and selective packaging of EVs remain poorly understood. Rab family guanosine triphosphates (GTPases) and their regulators are important membrane trafficking organizers. In this study, we investigated the role of select Rab GTPases on the regulation of EV release. We found that microRNAs target Rab GTPases to regulate EV release from lung cancer cell lines. In particular, Rab32 is a target of miR-124a, and siRNA and miRNA mediated inhibition of Rab32 leads to impaired EV secretion. The downstream implications for microRNA-based regulation of EV release are currently under investigation.

Extracellular vesicles (EVs) are heterogenous membrane-encapsulated vesicles secreted by every cell into the extracellular environment. EVs carry bioactive molecules, including proteins, lipids, DNA, and different RNA forms, which can be internalized by recipient cells, thus altering their biological characteristics. Given that EVs are commonly found in most body fluids, they have been widely described as mediators of communication in several physiological and pathological processes, including cancer. Moreover, their easy detection in biofluids makes them potentially useful candidates as tumor biomarkers. In this manuscript, we review the current knowledge regarding EVs and non-coding RNAs and their role as drivers of the metastatic process in lung cancer. Furthermore, we present the most recent applications for EVs and non-coding RNAs as cancer therapeutics and their relevance as clinical biomarkers.

A significant percentage of the general population reports gastrointestinal and non-gastrointestinal symptoms caused by wheat and/or gluten ingestion, even though they do not suffer from celiac disease (CD) or wheat allergy (WS), because they test negative both for CD-specific serology and histopathology. All patients report improvement of symptoms on a gluten-free diet. This clinical condition has been named non-celiac gluten sensitivity (NCGS). The objective of this paper was to review some studies regarding the pathogenesis of NCGS to summarize the current hypotheses about the mechanisms, which can lead to NCGS. Particular attention was given to the immunologic and the malabsorptive hypotheses. We reviewed data of our previous studies involving patients diagnosed with NCWS by means of double-blind placebo-controlled wheat challenge. The data indicating a possible wheat allergy diagnosis were examined and other data in the literature were reviewed; we also reviewed the putative role of fermentable oligosaccharides disaccharides monosaccharides and polyols (FODMAPs) and of α-amylasetrypsin inhibitors (ATIs) proteins in the NCGS pathogenesis. NCGS pathogenesis has been attributed to very different mechanisms, among others: i) activation of innate and adaptive immunity (the immune/allergic mechanisms) induced by gluten or the non-gluten ATI family; ii) incomplete digestion and/or absorption of FODMAPs (the intolerance mechanisms); and iii) psychological effect.We suggest that NCGS is a heterogeneous condition, which includes different subgroups of patients who have different pathogenic mechanisms: strong data suggest a direct pathogenic immunogenic/inflammatory role of wheat-cereal proteins (not only gluten) in a subgroup, probably the largest, of these patients.

Dyspnea is the main clinical manifestation of acute respiratory failure; however, it can be induced by different causes that require differential diagnosis. In elderly patients, the medical examination is limited by cognitive and physical deficits and the genesis of dyspnea is often multifactorial; therefore, the definition of the causes is more complex. In recent years, among the various diagnostic methods, pulmonary ultrasonography has found increasing interest and the main patterns of ultrasound semeiotics have been described, specially in the emergency and critical care setting, as for pneumonia, pleural effusion, pneumothorax, pulmonary infarction, interstitial pathology. The most recent Literature emphasizes the role of multi-organ ultrasound (pleuro-pulmonar; cardiac; caval and iliac-femoral venous), to diagnose with good accuracy the causes of dyspnea. In this review we report the lung ultrasound semeiotic patterns, useful for the bedside or pointof- care detection of the cause of dyspnea, particularly respiratory and/or cardiac, during the medical examination.

Over the last several decades, clinical evaluation and treatment of lung cancers have largely improved with the classification of genetic drivers of the disease, such as EGFR, ALK, and ROS1. There are numerous regulatory factors that exert cellular control over key oncogenic pathways involved in lung cancers. In particular, non-coding RNAs (ncRNAs) have a diversity of regulatory roles in lung cancers such that they have been shown to be involved in inducing proliferation, suppressing apoptotic pathways, increasing metastatic potential of cancer cells, and acquiring drug resistance. The dysregulation of various ncRNAs in human cancers has prompted preclinical studies examining the therapeutic potential of restoring and/or inhibiting these ncRNAs. Furthermore, ncRNAs demonstrate tissue-specific expression in addition to high stability within biological fluids. This makes them excellent candidates as cancer biomarkers. This review aims to discuss the relevance of ncRNAs in cancer pathology, diagnosis, and therapy, with a focus on lung cancer.

In the last 20 years, the functional roles for miRNAs in gene regulation have been well established. MiRNAs act as regulators in virtually all biological pathways and thus have been implicated in numerous diseases, including cancer. They are particularly relevant in regulating the basic hallmarks of cancer, including apoptosis, proliferation, migration, and invasion. Despite the substantial progress made in identifying the molecular mechanisms driving the deregulation of miRNAs in cancer, the clinical translation of these important molecules to therapy remains in its infancy. The paucity of vehicles available for the safe and efficient delivery of miRNAs and ongoing concerns for toxicity remain major obstacles to clinical application. Novel formulations and the development of new vectors have significantly improved the stability of oligonucleotides, increasing the effectiveness of therapy. Furthermore, the use of specific moieties for delivery in target tissues or cells has increased the specificity of treatment. The use of new technologies has allowed small but important steps toward more specific therapeutic delivery in tumor tissues and cells. Although a long road remains, the path ahead holds great potential. Currently, a few miRNA drugs are under investigation in human clinical trials with promising results ahead.

Transactive response DNA-binding protein 43 kDa (TDP-43, also known as TBPH in Drosophila melanogaster and TARDBP in mammals) is the main protein component of the pathological inclusions observed in neurons of patients affected by different neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) and fronto-temporal lobar degeneration (FTLD). The number of studies investigating the molecular mechanisms underlying neurodegeneration is constantly growing; however, the role played by TDP-43 in disease onset and progression is still unclear. A fundamental shortcoming that hampers progress is the lack of animal models showing aggregation of TDP-43 without overexpression. In this manuscript, we have extended our cellular model of aggregation to a transgenic Drosophila line. Our fly model is not based on the overexpression of a wild-type TDP-43 transgene. By contrast, we engineered a construct that includes only the specific TDP-43 amino acid sequences necessary to trigger aggregate formation and capable of trapping endogenous Drosophila TDP-43 into a non-functional insoluble form. Importantly, the resulting recombinant product lacks functional RNA recognition motifs (RRMs) and, thus, does not have specific TDP-43-physiological functions (i.e. splicing regulation ability) that might affect the animal phenotype per se. This novel Drosophila model exhibits an evident degenerative phenotype with reduced lifespan and early locomotion defects. Additionally, we show that important proteins involved in neuromuscular junction function, such as syntaxin (SYX), decrease their levels as a consequence of TDP-43 loss of function implying that the degenerative phenotype is a consequence of TDP-43 sequestration into the aggregates. Our data lend further support to the role of TDP-43 loss-of-function in the pathogenesis of neurodegenerative disorders. The novel transgenic Drosophila model presented in this study will help to gain further insight into the molecular mechanisms underlying neurodegeneration and will provide a valuable system to test potential therapeutic agents to counteract disease.

In the last two decades, RNA post-transcriptional modifications, including RNA editing, have been the subject of increasing interest among the scientific community. The efforts of the Human Genome Project combined with the development of new sequencing technologies and dedicated bioinformatic approaches created to detect and profile RNA transcripts have served to further our understanding of RNA editing. Investigators have determined that non-coding RNA (ncRNA) A-to-I editing is often deregulated in cancer. This discovery has led to an increased number of published studies in the field. However, the eventual clinical application for these findings remains a work in progress. In this review, we provide an overview of the ncRNA editing phenomenon in cancer. We discuss the bioinformatic strategies for RNA editing detection as well as the potential roles for ncRNA A to I editing in tumor immunity and as clinical biomarkers.

In packed low-moisture foods such as crackers, oxidation is generally the main cause of quality depletion during storage. It is commonly believed, but scarcely investigated, that product shelf life depends on the oxidative status of the lipid ingredients. In this study, the influence of oxidation degree of the ingredient sunflower oil on cracker oxidative stability and hence shelf life was investigated. To this aim, oil with increasing peroxide values (PVs) (5, 11, and 25 mEqO2/kgoil) was used to prepare crackers. Just after production, crackers presented similar peroxide and rancid odor intensity, probably due to the interactive pathways of oxidative and Maillard reactions. Crackers were packed and analyzed for PV and rancid odor during storage at 20, 40, and 60 °C. Rancid odor well discriminated cracker oxidative status. Relevant oxidation rates were used to develop a shelf life predictive model based on the peroxide value of the ingredient oil. It was estimated that an oil PV from 5 to 15 mEqO2/kgoil shortens cracker Shelf Life (SL) by 50%, independently of storage temperature. These results demonstrate the critical impact of ingredient quality on product performance on the market.

Nearly all classes of coding and non-coding RNA undergo post-transcriptional modification, as more than 150 distinct modification types have been reported. Since RNA modifications were first described over 50 years ago, our understanding of their functional relevance in cellular control mechanisms and phenotypes has truly progressed only in the last 15 years due to advancements in detection and experimental techniques. Specifically, the phenomenon of RNA methylation in the context of ncRNA has emerged as a novel process in the arena of epitranscriptomics. Methylated ncRNA molecules may indeed contribute to a potentially vast functional panorama, from regulation of post-transcriptional gene expression to adaptive cellular responses. Recent discoveries have uncovered novel dynamic mechanisms and new layers of complexity, paving the way to a greater understanding of the role of such phenomena within the broader molecular cellular context of human disease.

TDP-43 is an ubiquitous and highly conserved ribonucleoprotein involved in several cellular processes including pre-mRNA splicing, transcription, mRNA stability and transport. Notwithstanding the evidence of TDP-43 involvement in the pathogenesis of different neurodegenerative disorders (i.e. ALS and FTLD), the underlying mechanisms are still unclear. Given the high degree of functional similarity between the human and fly orthologs of TDP-43, Drosophila melanogaster is a simple and useful model to study the pathophysiological role of this protein in vivo. It has been demonstrated that the depletion of the TDP-43 fly ortholog (tbph) induces deficient locomotive behaviors and reduces life span and anatomical defects at the neuromuscular junction. In this study, using the known binding specificity of TDP-43/tbph for (UG) repeated sequences, we performed a bioinformatic screening for fly genes with at least 6 (TG) repeats in a row within the 3'-UTR regions in order to identify the genes that might be regulated by this factor. Among these genes, we were able to identify RhoGAPp190 as a potential target of the tbph-mediated neurodegeneration. RhoGAPp190 is a negative regulator of Drosophila RhoA, a GTPase protein implicated in the fine modulation of critical cellular processes including axon branch stability and motor axon defasciculation at muscle level and cognitive processes. We were able to demonstrate that the RhoGAPp190 expression is upregulated in a tbph-null fly model, providing evidence that this deregulation is associated to tbph silencing. Our results introduce RhoGAPp190 as a novel potential mediator in the complex scenario of events resulting from in vivo tbph loss-of-function.

Efficiency improvement is one of three patterns a public utility should follow in order to get funds for investments realization. The other two are recourse to bank loans or to private equity and tariff increase. Efficiency can be improved, for example, by growth and vertical integration and may be conditioned by environmental variables, such as customer and output density. Prior studies into the effects of these variables on the efficiency of water utilities do not agree on certain points (e.g., scale and economies of scope) and rarely consider others (e.g., density economies). This article aims to contribute to the literature by analysing the efficiency of water utilities in Denmark, observing the effects of operational and environmental variables. The method is based on two-stage Data Envelopment Analysis (DEA) applied to 101 water utilities. We found that the efficiency of the water sector was not affected by the observed variables, whereas that of wastewater was improved by smaller firm size, vertical integration strategy, and higher population density.

Alterations in TDP-43 are commonly found in patients suffering from amyotrophic lateral sclerosis (ALS) and the genetic suppression of the conserved homologue in Drosophila (TBPH) provokes alterations in the functional organization of motoneuron synaptic terminals, resulting in locomotive defects and reduced life span. To gain more insight into this pathological process, it is of fundamental importance to establish when during the fly life cycle the lack of TBPH affects motoneuron activity and whether this is a reversible phenomenon. To achieve this, we conditionally expressed the endogenous protein in TBPH minus Drosophila neurons and found that TBPH is a short lived protein permanently required for Drosophila motility and synaptic assembly through the direct modulation of vesicular proteins, such as Syntaxin 1A, indicating that synaptic transmission defects are early pathological consequences of TBPH dysfunction in vivo. Importantly, TBPH late induction is able to recover synaptogenesis and locomotion in adult flies revealing an unexpected late-stage functional and structural neuronal plasticity. These observations suggest that late therapeutic approaches based on TDP-43 functionality may also be successful for the human pathology.

The aim of this study was to estimate the determinants of residential water demand for chief towns of every Italian province, in the period 2007–2009, using the linear mixed-effects model estimated with the restricted-maximum-likelihood method. Results confirmed that the applied tariff had a negative effect on residential water consumption and that it was a relevant driver of domestic water consumption. Moreover, income per capita had a positive effect on water consumption. Among measured climatic and geographical features, precipitation and altitude exerted a strongly significant negative effect on water consumption, while temperature did not influence water demand. Further, data show that small towns in terms of population served were characterized by lower levels of consumption. Water utilities ownership itself did not have a significant effect on water consumption but tariffs were significantly lower and residential water consumption was higher in towns where the water service was managed by publicly owned water utilities. However, further research is needed to gain a better understanding of the connection between ownership of water utilities and water prices and water consumption.

The aim of this Special Issue is to gather evidence on the impact of price policies (PP) and non-price policies (NPP) in shaping residential water use in a context of increased water scarcity. Indeed, a large body of the empirical economic literature on residential water demand has been devoted to measuring the impact of PP (water price increases, use of block rate pricing or peak pricing, etc.). The consensus is that the residential water demand is inelastic with respect to water price, but not perfectly. Given the low water price elasticity, pricing schemes may not always be effective tools for modifying household water behaviors. This is puzzling since increasing the water price is still viewed by public authorities as the most direct economic tool for inducing water conservation behaviors. Additional evidence regarding the use of PP in shaping residential water use is then required. More recently, it has been argued that residential consumers may react to NPP, such as water conservation programs, education campaigns, or smart metering. NPP are based on the idea that residential water users can implement strategies that will result in water savings via changing their individual behaviors. Feedback information based on smart water metering is an example of approach used by some water utilities. There are still large gaps in the knowledge on the residential water demand, and in particular on the impact of PP and NPP on residential water use, household water affordability and water service performance. These topics are addressed in this Special Issue “Advances in the Economic Analysis of Residential Water Use”.

Over the past decade, evidence has identified a link between protein aggregation, RNA biology, and a subset of degenerative diseases. An important feature of these disorders is the cytoplasmic or nuclear aggregation of RNA-binding proteins (RBPs). Redistribution of RBPs, such as the human TAR DNA-binding 43 protein (TDP-43) from the nucleus to cytoplasmic inclusions is a pathological feature of several diseases. Indeed, sporadic and familial forms of amyotrophic lateral sclerosis (ALS) and fronto-temporal lobar degeneration share as hallmarks ubiquitin-positive inclusions. Recently, the wide spectrum of neurodegenerative diseases characterized by RBPs functions’ alteration and loss was collectively named proteinopathies. Here, we show that TBPH (TAR DNA-binding protein-43 homolog), the Drosophila ortholog of human TDP-43 TAR DNA-binding protein-43, interacts with the arcRNA hsrω and with hsrω-associated hnRNPs. Additionally, we found that the loss of the omega speckles remodeler ISWI (Imitation SWI) changes the TBPH sub-cellular localization to drive a TBPH cytoplasmic accumulation. Our results, hence, identify TBPH as a new component of omega speckles and highlight a role of chromatin remodelers in hnRNPs nuclear compartmentalization.

We evaluated in elderly subjects (a) the ability of GFR formulas to discriminate chronic kidney disease (CKD), (b) the correlation between renal morphology and function, and (c) the usefulness of combined r-US and GFR formulas to detect CKD. A total of 72 patients were enrolled (mean age 80 ± 7 years, male sex 44%, serum creatinine 0.98 ± 0.42 mg/dL, and CKD 57%). Cockcroft-Gault showed the highest sensitivity (78%) and specificity (94%) for CKD and was correlated with kidney volume (R=0.68, P

Glioblastoma multiforme (GBM) is one of the most deadly types of cancer. To date, the best clinical approach for treatment is based on administration of temozolomide (TMZ) in combination with radiotherapy. Much evidence suggests that the intracellular level of the alkylating enzyme O(6)-methylguanine-DNA methyltransferase (MGMT) impacts response to TMZ in GBM patients. MGMT expression is regulated by the methylation of its promoter. However, evidence indicates that this is not the only regulatory mechanism present. Here, we describe a hitherto unknown microRNA-mediated mechanism of MGMT expression regulation. We show that miR-221 and miR-222 are upregulated in GMB patients and that these paralogues target MGMT mRNA, inducing greater TMZ-mediated cell death. However, miR-221/miR-222 also increase DNA damage and, thus, chromosomal rearrangements. Indeed, miR-221 overexpression in glioma cells led to an increase in markers of DNA damage, an effect rescued by re-expression of MGMT. Thus, chronic miR-221/222-mediated MGMT downregulation may render cells unable to repair genetic damage. This, associated also to miR-221/222 oncogenic potential, may poor GBM prognosis.

Lung cancer is the leading cause of cancer mortality in the world today. Although some advances in lung cancer therapy have been made, patient survival is still poor. MicroRNAs (miRNAs) can act as oncogenes or tumor-suppressor genes in human malignancy. The miR-34 family consists of tumor-suppressive miRNAs, and its reduced expression has been reported in various cancers, including non-small cell lung cancer (NSCLC). In this study, we found that miR-34a and miR-34c target platelet-derived growth factor receptor alpha and beta (PDGFR-α and PDGFR-β), cell surface tyrosine kinase receptors that induce proliferation, migration and invasion in cancer. MiR-34a and miR-34c were downregulated in lung tumors compared to normal tissues. Moreover, we identified an inverse correlation between PDGFR-α/β and miR-34a/c expression in lung tumor samples. Finally, miR-34a/c overexpression or downregulation of PDGFR-α/β by siRNAs, strongly augmented the response to TNF-related apoptosis inducing ligand (TRAIL) while reducing migratory and invasive capacity of NSCLC cells.

TDP-43 is an evolutionarily conserved RNA binding protein recently associated with the pathogenesis of different neurological diseases. At the moment, neither its physiological role in vivo nor the mechanisms that may lead to neurodegeneration are well known. Previously, we have shown that TDP-43 mutant flies presented locomotive alterations and structural defects at the neuromuscular junctions. We have now investigated the functional mechanism leading to these phenotypes by screening several factors known to be important for synaptic growth or bouton formation. As a result we found that alterations in the organization of synaptic microtubules correlate with reduced protein levels in the microtubule associated protein futsch/MAP1B. Moreover, we observed that TDP-43 physically interacts with futsch mRNA and that its RNA binding capacity is required to prevent futsch down regulation and synaptic defects.

Akt is a serine threonine kinase with a major role in transducing survival signals and regulating proteins involved in apoptosis. To find new interactors of Akt involved in cell survival, we performed a two-hybrid screening in yeast using human full-length Akt c-DNA as bait and a murine c-DNA library as prey. Among the 80 clones obtained, two were identified as Bcl-w. Bcl-w is a member of the Bcl-2 family that is essential for the regulation of cellular survival, and that is up-regulated in different human tumors, such as gastric and colorectal carcinomas. Direct interaction of Bcl-w with Akt was confirmed by immunoprecipitation assays. Subsequently, we addressed the function of this interaction: by interfering with the activity or amount of Akt, we have demonstrated that Akt modulates the amount of Bcl-w protein. We have found that inhibition of Akt activity may promote apoptosis through the downregulation of Bcl-w protein and the consequential reduction in interaction of Bcl-w with pro-apoptotic members of the Bcl-2 family. Our data provide evidence that Bcl-w is a new member of the Akt pathway and that Akt may induce anti-apoptotic signals at least in part through the regulation of the amount and activity of Bcl-w.