Phase behavior and critical activated dynamics of limited-valence DNA nanostars

Colloidal particles with directional interactions are key in the realization of new colloidal materials with possibly unconventional phase behaviors. Here we exploit DNA self-assembly to produce bulk quantities of "DNA stars" with three or four sticky terminals, mimicking molecules with controlled limited valence. Solutions of such molecules exhibit a consolution curve with an upper critical point, whose temperature and concentration decrease with the valence. Upon approaching the critical point from high temperature, the intensity of the scattered light diverges with a power law, whereas the intensity time autocorrelation functions show a surprising two-step relaxation, somehow reminiscent of glassy materials. The slow relaxation time exhibits an Arrhenius behavior with no signs of criticality, demonstrating a unique scenario where the critical slowing down of the concentration fluctuations is subordinate to the large lifetime of the DNA bonds, with relevant analogies to critical dynamics in polymer solutions. The combination of equilibrium and dynamic behavior of DNA nanostars demonstrates the potential of DNA molecules in diversifying the pathways toward collective properties and self-assembled materials, beyond the range of phenomena accessible with ordinary molecular fluids

ROLE OF THE PRKCA GENE AND OF MICRORNAS IN THE SUSCEPTIBILITY TO MULTIPLE SCLEROSIS

Multiple sclerosis (MS) is a multifactorial neurological disorder characterized by chronic inflammation, demyelination, and axonal damage, probably caused by an altered immune response against myelin antigens. Despite extensive studies, the genetic and environmental determinants of MS are still poorly understood. In this thesis, we analyzed the role of the PRKCA gene that encodes the protein kinase C alpha, and has been involved in MS both by linkage and association studies in different populations (Finns, Canadians, and UK population), as well as the possible involvement of microRNAs (miRNAs) in the pathogenesis of MS. As far as PRKCA is concerned, a case-control association study performed on the Italian population allowed us to identify two association signals in the PRKCA gene: a protective allele mapping in the promoter region (P=0.033; OR=0.12, 95% CI=0.015-0.94) and a risk haplotype, partially overlapping the predisposing haplotypes observed both in Finns and Canadians within PRKCA intron 3 (P=7.4*10-4; OR=1.57, 95% CI=1.24-1.99). We identified and characterized, both in vitro and in vivo, the functional variants underlying these association signals, and demonstrated that higher PRKCA expression levels play a protective role towards MS predisposition. Moreover, we describe here for the first time so-far unknown PRKCA transcript isoforms, which are differentially expressed in MS cases and controls, and eventually lead to the synthesis of aberrant proteins, potentially dangerous for the cells. Furthermore, we characterized the MIR634 gene, mapping within the PRKCA locus, and demonstrated its involvement in the regulation of PRKCA expression. As far as miRNAs are concerned, we monitored the differential expression of several immunity-related miRNAs in peripheral blood mononuclear cells of MS patients and healthy controls, and identified miR-155 as the most upregulated miRNA in MS cases (fold change=3.30; P=0.013). Interestingly, this miRNA was previously reported to be up-regulated also in MS brain lesions. The role of miR-155 in MS susceptibility was also investigated by genotyping four single nucleotide polymorphisms mapping in the miR-155 genomic region. A three-SNP haplotype resulted associated with the disease status (P=0.035; OR=1.36, 95% CI=1.05-1.77), suggesting that this locus strongly deserves further investigations. In conclusion, this thesis work points to a possible role of post-transcriptional regulation mechanisms in the pathogenesis of MS, particularly supporting the uprising hypothesis that in MS a dysregulation of the splicing may play a pivotal role in MS

Modularization Techniques for Active Rules Design

ACM TOD

Differential expression of microRNAs in peripheral blood mononuclear cells of Multiple Sclerosis patients

MicroRNAs (miRNAs) are a class of short (~22 nucleotides) single-stranded RNAs that modulate the expression of multiple target mRNAs by inducing either translational repression or mRNA degradation. MiRNAs have emerged has key post-transcriptional regulators of diverse biological processes and their deregulation has been implicated in several complex human diseases, including neurodegenerative and inflammatory disorders. Multiple sclerosis (MS) is a multifactorial disease of the central nervous system characterized by chronic inflammation, demyelination, axonal damage, and progressive neurological dysfunction. In spite of extensive research, the molecular events involved in the initiation and progression of MS are still poorly understood. To be relevant for MS pathogenesis, candidate genes would be expected to be expressed either in tissues relevant for immune response, or in tissues affected by the disease process. Several miRNAs are known to be expressed specifically in the immune system and miRNA-dependent alterations in gene expression in hematopoietic cells are critical for mounting an appropriate immune response. Moreover, deregulation of hematopoietic-specific miRNA expression may result in defects in both central and peripheral tolerance. Therefore, we sought to explore the possible involvement of miRNAs in MS by monitoring for differential expression of specific miRNAs in peripheral blood mononuclear cells (PBMCs) of MS patients and healthy controls. We selected a set of 22 candidate miRNAs that are expressed in the immune system and/or are transcribed from previously reported MS susceptibility loci. The differential expression of candidate miRNAs in cases versus controls was evaluated using a microbead-based technology. In a pilot experiment, performed on PBMC RNA of relapsing-remitting MS patients and controls, 4 miRNAs resulted >3 folds up-regulated in MS vs controls, whereas only the miR-150 resulted down-regulated (2-fold decrease). Interestingly, two of the most up-regulated miRNAs, mir-155 and mir-146a, have been reported to be altered also in rheumatoid arthritis and systemic lupus erythematosus, suggesting shared pathogenic mechanisms between different chronic inflammatory diseases. This is, to our knowledge, the first evidence that the expression of specific miRNAs is altered in MS