Functional interplay between MyoD and CTCF in regulating long-range chromatin interactions during differentiation

Higher-order chromatin structures appear to be dynamically arranged during development and differentiation. However, the molecular mechanism underlying their maintenance or disruption and their functional relevance to gene regulation are poorly understood. We recently described a dynamic long-range chromatin interaction between the gene promoter of the cdk inhibitor p57(kip2) (also known as Cdkn1c) and the imprinting control region KvDMR1 in muscle cells. Here, we show that CTCF, the best characterized organizer of long-range chromatin interactions, binds to both the p57(kip2) promoter and KvDMR1 and is necessary for the maintenance of their physical contact. Moreover, we show that CTCF-mediated looping is required to prevent p57(kip2) expression before differentiation. Finally, we provide evidence that the induction of p57(kip2) during myogenesis involves the physical interaction of the muscle-regulatory factor MyoD with CTCF at KvDMR1, the displacement of the cohesin complex subunit Rad21 and the destabilization of the chromatin loop. The finding that MyoD affects chromatin looping at CTCF-binding sites represents the first evidence that a differentiation factor regulates chromatin-loop dynamics and provides a useful paradigm for gaining insights into the developmental regulation of long-range chromatin contacts. © 2014. Published by The Company of Biologists Ltd

Petroleum systems modelling as an exploration tool: from surface seismic acquisition to basin modelling: a case study from a periplatform basin in Northern Adriatic

The Adriatic Sea hosts important historically explored hydrocarbon provinces. In the northern areas, the main sources of hydrocarbon productivity are from biogenic gas reservoirs located in Quaternary thick deltaic depositional systems related to the Apennine and Dinaric foreland domain. In addition, Mesozoic thermogenic oil plays were identified and exploited in the Southern areas. Herein, we use basin and petroleum systems modelling (BPSM) techniques to investigate the still unproved thermogenic hydrocarbon potential of deep Mesozoic basins in the North Adriatic Sea and to correlate the results with the sedimentary evolution of proven petroleum systems in the Central and South Adriatic, both in the Italian and Croatian offshore areas. We used the regional CROP-M16 marine seismic profile in the eastern segment of the study area, where it cuts the western margin of the Dinaric carbonate platform and reaches the pelagic domain of the Umbria-Marche basin intersecting the Barbara isolated platform (Figure 1). We reprocessed the 2D seismic data set to obtain the correct dips and depths of the geological structures from a depth-migrated profile. During the interpretation phase, we integrated the information from profile CROP-M16 with the orthogonal CROP-M17B and C profiles that are intersecting the Alessandra-001 exploration well. The interpretation allowed a detailed analysis of the main lithological discontinuities and of the fault system. We used the interpreted features together with the petrophysical parameters inferred by the available wells in the area and by literature, for 2D petroleum systems modelling. Our simulation integrated the available information to produce a set of possible scenarios of basin development, with the objective to understand the maturation and migration of hydrocarbons

Regulation of p57KIP2 during muscle differentiation: role of Egr1, Sp1 and DNA hypomethylation

The cdk inhibitor p57(kip2) plays a critical role in many differentiation processes by performing not only redundant but also specific functions. Compared to other cdk inhibitors, p57(kip2) shows a more restricted expression pattern during development and in adult tissues. We have previously reported that in muscle cells, p57(kip2) is induced by the myogenic factor MyoD through an indirect mechanism involving p73 proteins as intermediaries. We have also reported that p57(kip2) shows a differential responsiveness to MyoD-dependent regulation in different cell types. In this work we have further investigated the molecular mechanism by which MyoD activates p57 promoter. We show that the minimal promoter element able to confer MyoD responsiveness contains multiple Sp1 and Egr1 recognition sites and that both transcription factors are necessary for the increase in p57 RNA. We also suggest that the role of MyoD-induced p73 consists in promoting the binding of Sp1 to p57(kip2) promoter. Moreover, we show that Egr1 and Sp1 are concomitantly recruited to p57 promoter in vivo only in differentiation conditions and only in responsive cells. Bisulfite sequencing suggested a functional link between the methylation status and the differential activity of p57 promoter, both during differentiation and in distinct cell types. These results, which highlight the involvement of epigenetic factors in the regulation of p57 expression in muscle cells, could be of general relevance to explain its tissue and cell type restriction during development

MyoD regulates p57kip2 expression by interacting with a distant cis-element and modifying a higher order chromatin structure

The bHLH transcription factor MyoD, the prototypical master regulator of differentiation, directs a complex program of gene expression during skeletal myogenesis. The up-regulation of the cdk inhibitor p57kip2 plays a critical role in coordinating differentiation and growth arrest during muscle development, as well as in other tissues. p57kip2 displays a highly specific expression pattern and is subject to a complex epigenetic control driving the imprinting of the paternal allele. However, the regulatory mechanisms governing its expression during development are still poorly understood. We have identified an unexpected mechanism by which MyoD regulates p57kip2 transcription in differentiating muscle cells. We show that the induction of p57kip2 requires MyoD binding to a long-distance element located within the imprinting control region KvDMR1 and the consequent release of a chromatin loop involving p57kip2 promoter. We also show that differentiation-dependent regulation of p57kip2, while involving a region implicated in the imprinting process, is distinct and hierarchically subordinated to the imprinting control. These findings highlight a novel mechanism, involving the modification of higher order chromatin structures, by which MyoD regulates gene expression. Our results also suggest that chromatin folding mediated by KvDMR1 could account for the highly restricted expression of p57kip2 during development and, possibly, for its aberrant silencing in some pathologies. © 2012 The Author(s)

Mutation in the VP1-LDV motif of the murine polyomavirus affects viral infectivity and conditions virus tissue tropism in vivo.

The first contact of a virus with the host cell surface and further entry are important steps for a successful outcome of the infection process and for the virus-associated pathogenicity. We have previously shown that the entry of the murine Polyomavirus (Py) into fibroblasts is a multi-step process involving, at least, the attachment to primary sialic acids (SA)-containing cell receptors followed by post-binding interaction with secondary receptors, such as the alpha4beta1 integrin, likely through the VP1-LDV motif. Here we report on the functional role of the VP1-LDV motif in Py infectivity and in vivo virus tissue tropism. For this purpose, we have characterized a recombinant virus mutant, PyLNV, harboring a single aa substitution in this motif (D138N). Although not critical for virus viability, the D138N substitution abrogates the post-attachment Py-alpha4beta1 interaction, rendering the PyLNV mutant virus twofold less infectious than the Py wild-type (Wt) in alpha4beta1-positive fibroblasts. To study the putative role of the VP1-LDV motif in vivo, newborn C57BL/6 mice were inoculated with PyWt or PyLNV and, after six days, organs were analyzed for the presence of viral DNA. Intriguingly, PyLNV showed an altered spectrum of in vivo replication compared with PyWt, particularly in the skin and in the kidney. The implication of Py-alpha4beta1 integrin interaction in conditioning tissue-specificity of virus replication is discussed.Journal ArticleResearch Support, Non-U.S. Gov'tinfo:eu-repo/semantics/publishe

The Importance of Knowledge Assets on University-Industry Collaboration: A Preliminary Analysis

Abstract. Intellectual capital is an important base for inter-organizational collaborative activities, including the industry-academic collaboration studied here. A preliminary study was conducted with analytic hierarchy process method, trying to know the dimensionalized associations between intellectual capital (i.e., Human, Structural, Customers, Organizational, Process, knowledge, and innovation capitals) and industry-academic collaboration (i.e., academic engagement and commercialization). Analyzing expert opinions, we gained matrices and priority scores, indicating that different forms of intellectual capital have different influences on academic engagement versus commercialization. The results remind scholarly works to look into detailed and differentiated mechanisms that utilize intellectual capital for governing industry-academic collaboration.Keywords. Intellectual capital, Industry-academic collaboration, Analytic hierarchy process; Taiwan.JEL. M10; L33; L52

Role of NFIB in normal hematopoiesis and MyeloproliferativeNeoplasms

Introduction: The three-canonical BCR-ABL negative myeloproliferative neoplasms (MPN), Polycythemia Vera (PV), Essential Thrombocytosis (ET) and Primary Myelofibrosis (PMF) are chronic diseases, which share the risk of disease evolution to an acute leukemia. In MPN, the most representative molecular lesion is a substitution of a valine for a phenylalanine (JAK2V617F) in the auto-inhibitory domain of JAK2, resulting in the constitutive expression of the gene.This point mutation has an incidence of about 96% in PV, 65% in PMF and 55% in ET. Microsatellite studies on chromosome 9 show the presence of a uniparental acquired disomy (UPD) of the short arm (9p), where JAK2 is located, as a common defect in MPN (Kralovics et al., ExpHematol. 2002). In a cohort study, all the samples with the 9pUPD were found positive for the JAK2V617F mutation (Klampfl et al, Blood 2011). The Nuclear Factor IB (NFIB) gene is present in the 9pUPD region, where a mutational hot spot takes place. NFIB belongs to the NFI family of CAAT box binding transcription factors, consisting of 4 separate genes (NFIA, -B, -C, -X). NFIA is a post-transcriptional target of myelopoiesis regulator miR-223, which plays a key role in directing the HSC/HPC maturation/differentiation into the erythroid or granulocytic lineages (Fazi et. al. Cell 2005; Starnes et al. Blood 2009). Genomic alterations of NFIA were detected in about 2% of MPN patients (Bernard, Leukemia 2009). The gene expression levels of NFIB are higher in CD34+HSC/HPC isolated from 5 JAK2V617F+PV patients than in normal controls (Berkofsky-Fessler, Clin Cancer Res 2010).However, the role of NFIB in normal and pathological hematopoiesis has not been yet investigated. Methods: DNA, mRNA, and proteins were isolated from human myeloid cell lines (K562, Hel and UKE-1) and buffy coat from peripheral blood (PB) and bone marrow (BM) cells isolated from MPN patients or healthy donors. Gene dosage and gene expression level were measured by qRT-PCR. Statistical analyses were used to calculate differences among and between groups by one way ANOVA and two-way t-test. Results: our preliminary data shows that: i) NFIB locus is amplified and its expression is increased in myeloid cell lines harboring the JAK2V617F; ii) NFIB is barely expressed in mononuclear cells isolated from healthy donors PB (n=23), while its expression increased in PB cells isolated from PV (n=18, p=0.034) and ET patients (n=27, p=0.005), independently from JAK2V617F status; iii) increased gene dosage of NFIB , paralleling JAK2 gene amplification is detected in MPN patients (n=27, p<0,001); iv) gene expression level of NFIB positively correlates with platelets count in ET patients (n=17, p=0.008). Conclusions: our preliminary data show the de-regulation of NFIB expression levels related to MPN, thus suggesting its role in MPN pathogenesis or disease evolution and usage as a marker for MPN diagnosis and/or prognosis

Demographic characteristics of study population and results of ANOVA analysis to compare each glaucoma severity subgroups and healthy controls.

<p>Demographic characteristics of study population and results of ANOVA analysis to compare each glaucoma severity subgroups and healthy controls.</p