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Maggi R.,Laboratory of Developmental Neuroendocrlnology | Maggi R.,Interunlverslty Center for the Research on the Molecular Bases of Reproductive Diseases | Zasso J.,Center for Integrative Biology | Conti L.,Center for Integrative Biology
Frontiers in Cellular Neuroscience | Year: 2015

The adult hypothalamus regulates many physiological functions and homeostatic loops, including growth, feeding and reproduction. In mammals, the hypothalamus derives from the ventral diencephalon where two distinct ventricular proliferative zones have been described. Although a set of transcription factors regulating the hypothalamic development has been identified, the exact molecular mechanisms that drive the differentiation of hypothalamic neural precursor cells (NPCs) toward specific neuroendocrine neuronal subtypes is yet not fully disclosed. Neurogenesis has been also reported in the adult hypothalamus at the level of specific niches located in the ventrolateral region of ventricle wall, where NPCs have been identified as radial glia-like tanycytes. Here we review the molecular and cellular systems proposed to support the neurogenic potential of developing and adult hypothalamic NPCs. We also report new insights on the mechanisms by which adult hypothalamic neurogenesis modulates key functions of this brain region. Finally, we discuss how environmental factors may modulate the adult hypothalamic neurogenic cascade. © 2015 Maggi, Zasso and Conti.


Gasperini L.,International School for Advanced Studies | Gasperini L.,Center for Integrative Biology | Meneghetti E.,International School for Advanced Studies | Legname G.,International School for Advanced Studies | And 2 more authors.
Frontiers in Neuroscience | Year: 2016

Essential elements as copper and iron modulate a wide range of physiological functions. Their metabolism is strictly regulated by cellular pathways, since dysregulation of metal homeostasis is responsible for many detrimental effects. Neurodegenerative disorders such as Alzheimer's disease, Parkinson's disease and prion diseases are characterized by alterations of metal ions. These neurodegenerative maladies involve proteins that bind metals and mediate their metabolism through not well-defined mechanisms. Prion protein, for instance, interacts with divalent cations via multiple metal-binding sites and it modulates several metal-dependent physiological functions, such as S-nitrosylation of NMDA receptors. In this work we focused on the effect of prion protein absence on copper and iron metabolism during development and adulthood. In particular, we investigated copper and iron functional values in serum and several organs such as liver, spleen, total brain and isolated hippocampus. Our results show that iron content is diminished in prion protein-null mouse serum, while it accumulates in liver and spleen. Our data suggest that these alterations can be due to impairments in copper-dependent cerulopalsmin activity which is known to affect iron mobilization. In prion protein-null mouse total brain and hippocampus, metal ion content shows a fluctuating trend, suggesting the presence of homeostatic compensatory mechanisms. However, copper and iron functional values are likely altered also in these two organs, as indicated by the modulation of metal-binding protein expression levels. Altogether, these results reveal that the absence of the cellular prion protein impairs copper metabolism and copper-dependent oxidase activity, with ensuing alteration of iron mobilization from cellular storage compartments. © 2016 Gasperini, Meneghetti, Legname and Benetti.


Petris G.,University of Trento | Casini A.,University of Trento | Montagna C.,University of Trento | Lorenzin F.,University of Trento | And 6 more authors.
Nature Communications | Year: 2017

In vivo application of the CRISPR-Cas9 technology is still limited by unwanted Cas9 genomic cleavages. Long-term expression of Cas9 increases the number of genomic loci non-specifically cleaved by the nuclease. Here we develop a Self-Limiting Cas9 circuit for Enhanced Safety and specificity (SLiCES) which consists of an expression unit for Streptococcus pyogenes Cas9 (SpCas9), a self-targeting sgRNA and a second sgRNA targeting a chosen genomic locus. The self-limiting circuit results in increased genome editing specificity by controlling Cas9 levels. For its in vivo utilization, we next integrate SLiCES into a lentiviral delivery system (lentiSLiCES) via circuit inhibition to achieve viral particle production. Upon delivery into target cells, the lentiSLiCES circuit switches on to edit the intended genomic locus while simultaneously stepping up its own neutralization through SpCas9 inactivation. By preserving target cells from residual nuclease activity, our hit and go system increases safety margins for genome editing.


Gallina L.,University of Bologna | Dal Pozzo F.,University of Bologna | Dal Pozzo F.,University of Liège | Galligioni V.,University of Bologna | And 3 more authors.
Antiviral Research | Year: 2011

Canine distemper virus (CDV) is a contagious and multisystemic viral disease that affects domestic and wild canines as well as other terrestrial and aquatic carnivores. The disease in dogs is often fatal and no specific antiviral therapy is currently available.In this study, we evaluated the in vitro antiviral activity against CDV of proanthocyanidin A2 (PA2), a phenolic dimer belonging to the class of condensed tannins present in plants. Our results showed that PA2 exerted in vitro antiviral activity against CDV with a higher selectivity index compared to ribavirin, included in our study for the previously tested anti-CDV activity. The time of addition assay led us to observe that PA2 was able to decrease the viral RNA synthesis and to reduce progeny virus liberation, at different times post infection suggesting multiple mechanisms of action including inhibition of viral replicative complex and modulation of the redox milieu. These data suggest that PA2, isolated from the bark of Aesculus hippocastanum, has potential usefulness as an anti-CDV compound inhibiting viral replication. © 2011 Elsevier B.V.


Ellen A.F.,Center for Integrative Biology | Ellen A.F.,University of Groningen | Zolghadr B.,University of Groningen | Zolghadr B.,Max Planck Institute for Terrestrial Microbiology | And 2 more authors.
Archaea | Year: 2010

Although archaea have a similar cellular organization as other prokaryotes, the lipid composition of their membranes and their cell surface is unique. Here we discuss recent developments in our understanding of the archaeal protein secretion mechanisms, the assembly of macromolecular cell surface structures, and the release of S-layer-coated vesicles from the archaeal membrane. © 2010 Albert F. Ellen et al.


Cereseto A.,Center for Integrative Biology | Giacca M.,International Center for Genetic Engineering and Biotechnology
Methods in Molecular Biology | Year: 2014

Advancements in fluorescent microscopy techniques now permit investigation of HIV-1 biology exploiting tools alternative to conventional molecular biology. Here we describe a novel, fluorescence-based method to visualize HIV-1 viral particles within intact nuclei of infected cells. This method allows investigating the localization of pre-integration complexes within the nuclear compartment with respect to the nuclear envelope and the chromatin territories. © 2014 Springer Science+Business Media, LLC.


Andreotti V.,Italian National Cancer Institute | Ciribilli Y.,Center for Integrative Biology | Monti P.,Italian National Cancer Institute | Bisio A.,Center for Integrative Biology | And 6 more authors.
PLoS ONE | Year: 2011

Background: The p53 tumor suppressor, which is altered in most cancers, is a sequence-specific transcription factor that is able to modulate the expression of many target genes and influence a variety of cellular pathways. Inactivation of the p53 pathway in cancer frequently occurs through the expression of mutant p53 protein. In tumors that retain wild type p53, the pathway can be altered by upstream modulators, particularly the p53 negative regulators MDM2 and MDM4. Methodology/Principal Findings: Given the many factors that might influence p53 function, including expression levels, mutations, cofactor proteins and small molecules, we expanded our previously described yeast-based system to provide the opportunity for efficient investigation of their individual and combined impacts in a miniaturized format. The system integrates i) variable expression of p53 proteins under the finely tunable GAL1,10 promoter, ii) single copy, chromosomally located p53-responsive and control luminescence reporters, iii) enhanced chemical uptake using modified ABC-transporters, iv) small-volume formats for treatment and dual-luciferase assays, and v) opportunities to co-express p53 with other cofactor proteins. This robust system can distinguish different levels of expression of WT and mutant p53 as well as interactions with MDM2 or 53BP1. Conclusions/Significance: We found that the small molecules Nutlin and RITA could both relieve the MDM2-dependent inhibition of WT p53 transactivation function, while only RITA could impact p53/53BP1 functional interactions. PRIMA-1 was ineffective in modifying the transactivation capacity of WT p53 and missense p53 mutations. This dual-luciferase assay can, therefore, provide a high-throughput assessment tool for investigating a matrix of factors that can influence the p53 network, including the effectiveness of newly developed small molecules, on WT and tumor-associated p53 mutants as well as interacting proteins.


Lecca P.,University of Trento | Lecca P.,Association for Computing Machinery | Re A.,Center for Integrative Biology
Lecture Notes in Computer Science (including subseries Lecture Notes in Artificial Intelligence and Lecture Notes in Bioinformatics) | Year: 2016

While computational systems biology provides a rich array of methods for network clustering, most of them are not suitable to capture cellular network dynamics. In the most common setting, computational algorithms seek to integrate the static information embedded in near-global interaction networks with the temporal information provided by time series experiments. We present a novel technique for temporally informed network module detection, named TD-WGcluster (Time Delay Weighted Graph CLUSTERing). TD-WGcluster utilizes four steps: (i) time-lagged correlations are calculated between any couple of interacting nodes in the network; (ii) an unsupervised version of k-means algorithm detects sub-graphs with similar time-lagged correlation; (iii) a fast-greedy optimization algorithm identify connected components by sub-graph; (iv) a geometric entropy is computed for each connected component as a measure of its complexity. TD-WGcluster notable feature is the attempt to account for temporal delays in the formation of regulatory modules during signal propagation in a network. © Springer International Publishing Switzerland 2016.


Mazza T.,Mendel Institute | Ballarini P.,École Centrale Paris | Guido R.,University of Calabria | Prandi D.,Center for Integrative Biology
IEEE/ACM Transactions on Computational Biology and Bioinformatics | Year: 2012

Important achievements in traditional biology have deepened the knowledge about living systems leading to an extensive identification of parts-list of the cell as well as of the interactions among biochemical species responsible for cell's regulation. Such an expanding knowledge also introduces new issues. For example, the increasing comprehension of the interdependencies between pathways (pathways cross-talk) has resulted, on one hand, in the growth of informational complexity, on the other, in a strong lack of information coherence. The overall grand challenge remains unchanged: to be able to assemble the knowledge of every "piece of a system in order to figure out the behavior of the whole (integrative approach). In light of these considerations, high performance computing plays a fundamental role in the context of in-silico biology. Stochastic simulation is a renowned analysis tool, which, although widely used, is subject to stringent computational requirements, in particular when dealing with heterogeneous and high dimensional systems. Here, we introduce and discuss a methodology aimed at alleviating the burden of simulating complex biological networks. Such a method, which springs from graph theory, is based on the principle of fragmenting the computational space of a simulation trace and delegating the computation of fragments to a number of parallel processes. © 2012 IEEE.


Xu Z.,New York University | Chen H.,New York University | Chen H.,Princeton University | Ling J.,New York University | And 5 more authors.
Genes and Development | Year: 2014

In vivo cross-linking studies suggest that the Drosophila transcription factor Bicoid (Bcd) binds to several thousand sites during early embryogenesis, but it is not clear how many of these binding events are functionally important. In contrast, reporter gene studies have identified >60 Bcd-dependent enhancers, all of which contain clusters of the consensus binding sequence TAATCC. These studies also identified clusters of TAATCC motifs (inactive fragments) that failed to drive Bcd-dependent activation. In general, active fragments showed higher levels of Bcd binding in vivo and were enriched in predicted binding sites for the ubiquitous maternal protein Zelda (Zld). Here we tested the role of Zld in Bcd-mediated binding and transcription. Removal of Zld function and mutations in Zld sites caused significant reductions in Bcd binding to known enhancers and variable effects on the activation and spatial positioning of Bcd-dependent expression patterns. Also, insertion of Zld sites converted one of six inactive fragments into a Bcd-responsive enhancer. Genome-wide binding experiments in zld mutants showed variable effects on Bcd-binding peaks, ranging from strong reductions to significantly enhanced levels of binding. Increases in Bcd binding caused the precocious Bcd-dependent activation of genes that are normally not expressed in early embryos, suggesting that Zld controls the genome-wide binding profile of Bcd at the qualitative level and is critical for selecting target genes for activation in the early embryo. These results underscore the importance of combinatorial binding in enhancer function and provide data that will help predict regulatory activities based on DNA sequence. © 2014 Xu et al.

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