A structural view of microRNA-target recognition

It is well established that the correct identification of the messenger RNA targeted by a given microRNA (miRNA) is a difficult problem, and that available methods all suffer from low specificity. We hypothesize that the correct identification of the pairing should take into account the effect of the Argonaute protein (AGO), an essential catalyst of the recognition process. Therefore, we developed a strategy named MiREN for building and scoring three-dimensional models of the ternary complex formed by AGO, a miRNA and 22 nt of a target mRNA that putatively interacts with it. We show here that MiREN can be used to assess the likelihood that an RNA molecule is the target of a given miRNA and that this approach is more accurate than other existing methods, usually based on sequence or sequence-related features. Our results also suggest that AGO plays a relevant role in the selection of the miRNA targets. Our method can represent an additional step for refining predictions made by faster but less accurate classical methods for the identification of miRNA targets

identification of molecular targets in the human genome

The purpose of the present study is to contribute to the field of functional genomics by developing, testing and applying computational methods to the problem of the evaluation of the effects of environmental and pharmacological molecules on genome expression. Original results are described in four independent sections The first two sections are devoted to the investigation of the coding potential of alternative splicing products in the human genome. Sections three and four are devoted to the application of computational techniques to investigate the molecular targets and the effects on their expression of molecules known to interfere with the physiological functions of a cell. In particular these techniques were applied on a class of compounds (tocotrienols) constituents of the Vitamin E

Structural and biochemical insights of CypA and AIF interaction

The Cyclophilin A (CypA)/Apoptosis Inducing Factor (AIF) complex is implicated in the DNA degradation in response to various cellular stress conditions, such as oxidative stress, cerebral hypoxia-ischemia and traumatic brain injury. The pro-apoptotic form of AIF (AIF(Δ1-121)) mainly interacts with CypA through the amino acid region 370-394. The AIF(370-394) synthetic peptide inhibits complex formation in vitro by binding to CypA and exerts neuroprotection in a model of glutamate-mediated oxidative stress. Here, the binding site of AIF(Δ1-121) and AIF(370-394) on CypA has been mapped by NMR spectroscopy and biochemical studies, and a molecular model of the complex has been proposed. We show that AIF(370-394) interacts with CypA on the same surface recognized by AIF(Δ1-121) protein and that the region is very close to the CypA catalytic pocket. Such region partially overlaps with the binding site of cyclosporin A (CsA), the strongest catalytic inhibitor of CypA. Our data point toward distinct CypA structural determinants governing the inhibitor selectivity and the differential biological effects of AIF and CsA, and provide new structural insights for designing CypA/AIF selective inhibitors with therapeutic relevance in neurodegenerative diseases

Superconducting tunable flux qubit with direct readout scheme

We describe a simple and efficient scheme for the readout of a tunable flux qubit, and present preliminary experimental tests for the preparation, manipulation and final readout of the qubit state, performed in incoherent regime at liquid Helium temperature. The tunable flux qubit is realized by a double SQUID with an extra Josephson junction inserted in the large superconducting loop, and the readout is performed by applying a current ramp to the junction and recording the value for which there is a voltage response, depending on the qubit state. This preliminary work indicates the feasibility and efficiency of the scheme.Comment: 10 pages, 5 figure

Quality traits of saffron produced in Italy: geographical area effect and good practices

Saffron (Crocus sativus L.) is the most expensive spice in the world and is used in food, cosmetic and dyeing industries. Considering that the production of saffron is increasingly widespread in medium-small Italian farms as well as the scarceness of information and studies regarding the quality of the saffron produced in Italy, the principal aim of this study was to investigate the quality of Italian saffron. Qualitative analysis was conducted in accordance with ISO 3632 1,2:2010-2011 considering 484 samples collected over four years (2015-2018). In particular, moisture content, aroma strength (safranal), colouring strength (crocin) and flavour strength (picrocrocin) were assessed for each sample, and whether spice quality varied according to the geographical area where the spice was produced was also investigated. Qualitative analysis showed that the majority (84-93%) of the samples analysed are of the first quality category, regardless of the year of production. Moisture content and colouring strength are the factors that influence the quality of the spice most. Principal component analysis showed that quality is not influenced by the geographical area where the spice was produced. Finally, some best agricultural practices to obtain a high quality saffron spice are reported

Negative Feedback Regulation of Auxin Signaling by ATHB8/ACL5–BUD2 Transcription Module

ABSTRACT The role of auxin as main regulator of vascular differentiation is well established, and a direct correlation between the rate of xylem differentiation and the amount of auxin reaching the (pro)cambial cells has been proposed. It has been suggested that thermospermine produced by ACAULIS5 (ACL5) and BUSHY AND DWARF2 (BUD2) is one of the factors downstream to auxin contributing to the regulation of this process in Arabidopsis . Here, we provide an in-depth characterization of the mechanism through which ACL5 modulates xylem differentiation. We show that an increased level of ACL5 slows down xylem differentiation by negatively affecting the expression of homeodomain-leucine zipper ( HD–ZIP ) III and key auxin signaling genes. This mechanism involves the positive regulation of thermospermine biosynthesis by the HD–ZIP III protein ARABIDOPSIS THALIANA HOMEOBOX8 tightly controlling the expression of ACL5 and BUD2 . In addition, we show that the HD–ZIP III protein REVOLUTA contributes to the increased leaf vascularization and long hypocotyl phenotype of acl5 likely by a direct regulation of auxin signaling genes such as LIKE AUXIN RESISTANT2 ( LAX2 ) and LAX3 . We propose that proper formation and differentiation of xylem depend on a balance between positive and negative feedback loops operating through HD–ZIP III genes

Systems Biology Approaches for the Improvement of Oncolytic Virus-Based Immunotherapies

Oncolytic virus (OV)-based immunotherapy is mainly dependent on establishing an efficient cell-mediated antitumor immunity. OV-mediated antitumor immunity elicits a renewed antitumor reactivity, stimulating a T-cell response against tumor-associated antigens (TAAs) and recruiting natural killer cells within the tumor microenvironment (TME). Despite the fact that OVs are unspecific cancer vaccine platforms, to further enhance antitumor immunity, it is crucial to identify the potentially immunogenic T-cell restricted TAAs, the main key orchestrators in evoking a specific and durable cytotoxic T-cell response. Today, innovative approaches derived from systems biology are exploited to improve target discovery in several types of cancer and to identify the MHC-I and II restricted peptide repertoire recognized by T-cells. Using specific computation pipelines, it is possible to select the best tumor peptide candidates that can be efficiently vectorized and delivered by numerous OV-based platforms, in order to reinforce anticancer immune responses. Beyond the identification of TAAs, system biology can also support the engineering of OVs with improved oncotropism to reduce toxicity and maintain a sufficient portion of the wild-type virus virulence. Finally, these technologies can also pave the way towards a more rational design of armed OVs where a transgene of interest can be delivered to TME to develop an intratumoral gene therapy to enhance specific immune stimuli

Transcriptome analysis of human primary endothelial cells (HUVEC) from umbilical cords of gestational diabetic mothers reveals candidate sites for an epigenetic modulation of specific gene expression.

Within the complex pathological picture associated to diabetes, high glucose (HG) has ". per se" effects on cells and tissues that involve epigenetic reprogramming of gene expression. In fetal tissues, epigenetic changes occur genome-wide and are believed to induce specific long term effects. Human umbilical vein endothelial cells (HUVEC) obtained at delivery from gestational diabetic women were used to study the transcriptomic effects of chronic hyperglycemia in fetal vascular cells using Affymetrix microarrays. In spite of the small number of samples analyzed (n=6), genes related to insulin sensing and extracellular matrix reorganization were found significantly affected by HG. Quantitative PCR analysis of gene promoters identified a significant differential DNA methylation in TGFB2. Use of Ea.hy926 endothelial cells confirms data on HUVEC. Our study corroborates recent evidences suggesting that epigenetic reprogramming of gene expression occurs with persistent HG and provides a background for future investigations addressing genomic consequences of chronic HG. © 2014 Elsevier Inc

Molecular detection of Theileria equi in donkeys (Equus asinus) in a selected site in central Italy

Equine piroplasmosis is among the most relevant tick-borne diseases of domestic and wild equids. Donkeys (Equus asinus) represent a potential reservoir for haemoparasites by harbouring tick-transmitted haemoparasites that can infect horses. We investigated the occurrence of Babesia caballi and Theileria equi in a donkey farm in the province of Grosseto (central Italy) to determine their prevalence of infection. For this purpose, conventional polymerase chain reaction (PCR) assays were carried out on blood samples from 109 donkeys. These included 85 females and 24 males as well as 36 young, 49 adult and 24 old animals. B. caballi and T. equi were detected by using primers that amplify an approximately 560 bp portion of the small-subunit ribosomal DNA of most Babesia and Theileria species. All PCR-positive samples were sequenced to determine the species of amplified Babesia and Theileria DNA. Sequencing data analysis revealed that 36 (33%, 95% CI: 24.2-40.9%) donkeys were positive for T. equi DNA. No samples were positive for B. caballi DNA. T. equi PCR-positivity drastically increased with age (from 0% to 46.9% and 54.2%) and was not significantly associated with the gender. These results highlight the high molecular prevalence of T. equi in a donkey farm of central Italy and support the role of donkeys as carriers and reservoirs of theileriosis for horses. The lack of B. caballi DNA needs further investigation

MAISTAS: a tool for automatic structural evaluation of alternative splicing products

Motivation: Analysis of the human genome revealed that the amount of transcribed sequence is an order of magnitude greater than the number of predicted and well-characterized genes. A sizeable fraction of these transcripts is related to alternatively spliced forms of known protein coding genes. Inspection of the alternatively spliced transcripts identified in the pilot phase of the ENCODE project has clearly shown that often their structure might substantially differ from that of other isoforms of the same gene, and therefore that they might perform unrelated functions, or that they might even not correspond to a functional protein. Identifying these cases is obviously relevant for the functional assignment of gene products and for the interpretation of the effect of variations in the corresponding proteins