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Mannini L.,CNR Institute of Biomedical Technologies | Menga S.,CNR Institute of Biomedical Technologies | Musio A.,CNR Institute of Biomedical Technologies | Musio A.,Instituto Toscano Tumori
Human Mutation | Year: 2010

Cohesin is responsible for sister chromatid cohesion, ensuring the correct chromosome segregation. Beyond this role, cohesin and regulatory cohesin genes seem to play a role in preserving genome stability and gene transcription regulation. DNA damage is thought to be a major culprit for many human diseases, including cancer. Our present knowledge of the molecular basis underlying genome instability is extremely limited. Mutations in cohesin genes cause human diseases such as Cornelia de Lange syndrome and Roberts syndrome/SC phocomelia, and all the cell lines derived from affected patients show genome instability. Cohesin mutations have also been identified in colorectal cancer. Here, we will discuss the human disorders caused by alterations of cohesin function, with emphasis on the emerging role of cohesin as a genome stability caretaker. © 2010 Wiley-Liss, Inc.


MacIag A.,University of Milan | Peano C.,CNR Institute of Biomedical Technologies | Pietrelli A.,CNR Institute of Biomedical Technologies | Egli T.,Eawag - Swiss Federal Institute of Aquatic Science and Technology | And 2 more authors.
Nucleic Acids Research | Year: 2011

Specific promoter recognition by bacterial RNA polymerase is mediated by σ subunits, which assemble with RNA polymerase core enzyme (E) during transcription initiation. However, σ 70 (the housekeeping σ subunit) and σ S (an alternative σ subunit mostly active during slow growth) recognize almost identical promoter sequences, thus raising the question of how promoter selectivity is achieved in the bacterial cell. To identify novel sequence determinants for selective promoter recognition, we performed run-off/microarray (ROMA) experiments with RNA polymerase saturated either with σ 70 (Eσ 70) or with σ S (Eσ S) using the whole Escherichia coli genome as DNA template. We found that Eσ 70, in the absence of any additional transcription factor, preferentially transcribes genes associated with fast growth (e.g. ribosomal operons). In contrast, Eσ S efficiently transcribes genes involved in stress responses, secondary metabolism as well as RNAs from intergenic regions with yet-unknown function. Promoter sequence comparison suggests that, in addition to different conservation of the-35 sequence and of the UP element, selective promoter recognition by either form of RNA polymerase can be affected by the A/T content in the-10/+1 region. Indeed, site-directed mutagenesis experiments confirmed that an A/T bias in the-10/+1 region could improve promoter recognition by Eσ S. © 2011 The Author(s).


Ricchelli F.,CNR Institute of Biomedical Technologies | Sileikyte J.,University of Padua | Bernardi P.,University of Padua
Biochimica et Biophysica Acta - Bioenergetics | Year: 2011

The mitochondrial permeability transition is an increase of permeability of the inner mitochondrial membrane to ions and solutes with an exclusion size of about 1500 Da. It is generally accepted that the permeability transition is due to opening of a high-conductance channel, the permeability transition pore. Although the molecular nature of the permeability transition pore remains undefined, a great deal is known about its regulation and role in pathophysiology. This review specifically covers the characterization of the permeability transition pore by chemical modification of specific residues through photoirradiation of mitochondria after treatment with porphyrins. The review also illustrates the basic principles of the photodynamic effect and the mechanisms of phototoxicity and discusses the unique properties of singlet oxygen generated by specific porphyrins in discrete mitochondrial domains. These experiments provided remarkable information on the role, interactions and topology of His and Cys residues in permeability transition pore modulation and defined an important role for the outer membrane 18 kDa translocator protein (formerly known as the peripheral benzodiazepine receptor) in regulation of the permeability transition. © 2011 Elsevier B.V.


Minoia S.,Technical University Valencia Consejo Superior Of Investigaciones Cientificas | Carbonell A.,Donald Danforth Plant Science Center | Di Serio F.,CNR Plant Protection Institute | Gisel A.,CNR Institute of Biomedical Technologies | And 3 more authors.
Journal of Virology | Year: 2014

The identification of viroid-derived small RNAs (vd-sRNAs) of 21 to 24 nucleotides (nt) in plants infected by viroids (infectious non-protein-coding RNAs of just 250 to 400 nt) supports their targeting by Dicer-like enzymes, the first host RNA-silencing barrier. However, whether viroids, like RNA viruses, are also targeted by the RNA-induced silencing complex (RISC) remains controversial. At the RISC core is one Argonaute (AGO) protein that, guided by endogenous or viral sRNAs, targets complementary RNAs. To examine whether AGO proteins also load vd-sRNAs, leaves of Nicotiana benthamiana infected by potato spindle tuber viroid (PSTVd) were agroinfiltrated with plasmids expressing epitope-tagged versions of AGO1, AGO2, AGO3, AGO4, AGO5, AGO6, AGO7, AGO9, and AGO10 from Arabidopsis thaliana. Immunoprecipitation analyses of the agroinfiltrated halos revealed that all AGOs except AGO6, AGO7, and AGO10 associated with vd-sRNAs: AGO1, AGO2, and AGO3 preferentially with those of 21 and 22 nt, while AGO4, AGO5, and AGO9 additionally bound those of 24 nt. Deep-sequencing analyses showed that sorting of vd-sRNAs into AGO1, AGO2, AGO4, and AGO5 depended essentially on their 5′-terminal nucleotides, with the profiles of the corresponding AGO-loaded vd-sRNAs adopting specific hot spot distributions along the viroid genome. Furthermore, agroexpression of AGO1, AGO2, AGO4, and AGO5 on PSTVd-infected tissue attenuated the level of the genomic RNAs, suggesting that they, or their precursors, are RISC targeted. In contrast to RNA viruses, PSTVd infection of N. benthamiana did not affect miR168-mediated regulation of the endogenous AGO1, which loaded vd-sRNAs with specificity similar to that of its A. thaliana counterpart. © 2014, American Society for Microbiology.


Granzotto A.,CNR Institute of Biomedical Technologies | Zatta P.,CNR Institute of Biomedical Technologies
PLoS ONE | Year: 2011

It has been recently suggested that resveratrol can be effective in slowing down Alzheimer's disease (AD) development. As reported in many biochemical studies, resveratrol seems to exert its neuro-protective role through inhibition of β-amyloid aggregation (Aβ), by scavenging oxidants and exerting anti-inflammatory activities. In this paper, we demonstrate that resveratrol is cytoprotective in human neuroblastoma cells exposed to Aβ and or to Aβ-metal complex. Our findings suggest that resveratrol acts not through anti-aggregative pathways but mainly via its scavenging properties. © 2011 Granzotto, Zatta.


Castellana S.,University of Bari | Vicario S.,CNR Institute of Biomedical Technologies | Saccone C.,CNR Institute of Biomedical Technologies | Saccone C.,University of Bari
Genome Biology and Evolution | Year: 2011

The mitochondrial genome is a fundamental component of the eukaryotic domain of life, encoding for several important subunits of the respiratory chain, the main energy production system in cells. The processes by means of which mitochondrial DNA (mtDNA) replicates, expresses itself and evolves have been explored over the years, although various aspects are still debated. In this review, we present several key points in modern research on the role of evolutionary forces in affecting mitochondrial genomes in Metazoa. In particular, we assemble the main data on their evolution, describing the contributions of mutational pressure, purifying, and adaptive selection, and how they are related. We also provide data on the evolutionary fate of the mitochondrial synonymous variation, related to the nonsynonymous variation, in comparison with the pattern detected in the nucleus. Elevated mutational pressure characterizes the evolution of the mitochondrial synonymous variation, whereas purging selection, physiologically due to phenomena such as cell atresia and intracellular mtDNA selection, guarantees coding sequence functionality. This enables mitochondrial adaptive mutations to emerge and fix in the population, promoting mitonuclear coevolution. © The Author(s) 2010.


Navarro B.,CNR Institute of Plant virology | Gisel A.,CNR Institute of Biomedical Technologies | Rodio M.E.,CNR Institute of Plant virology | Delgado S.,Polytechnic University of Valencia | And 2 more authors.
Plant Journal | Year: 2012

How viroids, tiny non-protein-coding RNAs (∼250-400 nt), incite disease is unclear. One hypothesis is that viroid-derived small RNAs (vd-sRNAs; 21-24 nt) resulting from the host defensive response, via RNA silencing, may target for cleavage cell mRNAs and trigger a signal cascade, eventually leading to symptoms. Peach latent mosaic viroid (PLMVd), a chloroplast-replicating viroid, is particularly appropriate to tackle this question because it induces an albinism (peach calico, PC) strictly associated with variants containing a specific 12-14-nt hairpin insertion. By dissecting albino and green leaf sectors of Prunus persica (peach) seedlings inoculated with PLMVd natural and artificial variants, and cloning their progeny, we have established that the hairpin insertion sequence is involved in PC. Furthermore, using deep sequencing, semi-quantitative RT-PCR and RNA ligase-mediated rapid amplification of cDNA ends (RACE), we have determined that two PLMVd-sRNAs containing the PC-associated insertion (PC-sRNA8a and PC-sRNA8b) target for cleavage the mRNA encoding the chloroplastic heat-shock protein 90 (cHSP90), thus implicating RNA silencing in the modulation of host gene expression by a viroid. Chloroplast malformations previously reported in PC-expressing tissues are consistent with the downregulation of cHSP90, which participates in chloroplast biogenesis and plastid-to-nucleus signal transduction in Arabidopsis. Besides PC-sRNA8a and PC-sRNA8b, both deriving from the less-abundant PLMVd (-) strand, we have identified other PLMVd-sRNAs potentially targeting peach mRNAs. These results also suggest that sRNAs derived from other PLMVd regions may downregulate additional peach genes, ultimately resulting in other symptoms or in a more favorable host environment for viroid infection. © 2012 Blackwell Publishing Ltd.


Chiappori F.,CNR Institute of Biomedical Technologies | Merelli I.,CNR Institute of Biomedical Technologies | Colombo G.,CNR Institute of Chemistry of Molecular Recognition | Milanesi L.,CNR Institute of Biomedical Technologies | Morra G.,CNR Institute of Chemistry of Molecular Recognition
PLoS Computational Biology | Year: 2012

Investigating ligand-regulated allosteric coupling between protein domains is fundamental to understand cell-life regulation. The Hsp70 family of chaperones represents an example of proteins in which ATP binding and hydrolysis at the Nucleotide Binding Domain (NBD) modulate substrate recognition at the Substrate Binding Domain (SBD). Herein, a comparative analysis of an allosteric (Hsp70-DnaK) and a non-allosteric structural homolog (Hsp110-Sse1) of the Hsp70 family is carried out through molecular dynamics simulations, starting from different conformations and ligand-states. Analysis of ligand-dependent modulation of internal fluctuations and local deformation patterns highlights the structural and dynamical changes occurring at residue level upon ATP-ADP exchange, which are connected to the conformational transition between closed and open structures. By identifying the dynamically responsive protein regions and specific cross-domain hydrogen-bonding patterns that differentiate Hsp70 from Hsp110 as a function of the nucleotide, we propose a molecular mechanism for the allosteric signal propagation of the ATP-encoded conformational signal. © 2012 Chiappori et al.


Correa Leite M.L.,CNR Institute of Biomedical Technologies
Clinical and Applied Thrombosis/Hemostasis | Year: 2011

Aim: To examine the relationship between some blood parameters and mild kidney dysfunction. Participants and Methods: A total of 719 Italian men aged 42 to 74 years from a population-based survey carried out in the town of Bollate (Milan). General linear models were used to examine the variations in plasma fibrinogen, hematocrit, platelet counts, mean platelet volume, and uric acid across levels of kidney function (estimated on the basis of glomerular filtration rate [GFR]), adjusting for age, education, smoking, alcohol consumption, physical activity (evaluated as TV watching, engaging in sport practice, and walking/cycling), waist circumference, arm muscle area, high-density lipoprotein (HDL)-cholesterol, triglycerides, hypertension, diabetes, cardiovascular disease history, and nonsteroid anti-inflammatory, diuretic, and antihypertensive drug use. Results: Plasma fibrinogen and hematocrit levels increased, and platelet counts and mean platelet volume significantly decreased as GFR fell to <80 or <70 mL/min per 1.73 m 2; stratified analysis revealed an association with serum uric acid levels. Alterations compatible with an increased cardiovascular risk were particularly evident among the participants with higher uric acid levels, whereas those indicative of platelet dysfunction were found among participants with lower levels. Conclusions: Parameters affecting hemostasis and blood viscosity are altered when kidney function is only slightly reduced, and the patterns of these relationships seem to be influenced by the levels of serum uric acid, whose easy and inexpensive measurement could have prognostic value. © The Author(s) 2011.


Rizzi E.,CNR Institute of Biomedical Technologies
Methods in Molecular Biology | Year: 2015

The pyrosequencing methodology was applied in 2005 by 454 Lifesciences to the emerging field of next generation sequencing (NGS), revolutionizing the way of DNA sequencing. In the last years the same strategy grew up and was technologically updated, reaching a high throughput in terms of amount of generated sequences (reads) per run and in terms of length of sequence up to values of 800–1,000 bases. These features of pyrosequencing perfectly fit to bacterial genome sequencing for the de novo assemblies and resequencing as well. The approaches of shotgun and paired ends sequencing allow the bacterial genome finishing providing a high-quality data in few days with unprecedented results. © Springer Science+Business Media New York 2015.

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