125,277 research outputs found
3d combo chrrna–dna–immunofish
No full textEpigenetic mechanisms govern the quality, the stability, and the responsiveness of transcriptional programs to the environment. This regulation is ensured via the concerted action of different players (transcription factors, “reader” and “writer” enzymes, histone marks, structural proteins, noncoding regulatory RNAs) that flow in the 3D organization of the genome. Indeed, nuclear architecture participates in the punctual and cell-type-specific regulation of transcription. Hence, the fine dissection of these mechanisms will allow a deeper understanding of the gene expression machinery. In this chapter, we propose a challenging imaging-based method to study the reciprocal interactions between chromatin-associated RNAs, genomic loci, and chromatin compartment with a procedure of 3D COMBO chrRNA–DNA–ImmunoFISH, specifically developed to preserve the nuclear integrity and topology of human primary T cells. We believe that our protocol will contribute to the improvement of epigenetic studies on the 3D nuclear structure of T cell subsets, possibly shedding light on the still hidden epigenetic players responsible for the great plasticity and functional diversification exerted by T cells
Chromosome conformation capture in primary human cells
No full text3D organization of the genome, its structural and regulatory function of cell identity, is acquiring prominent features in epigenetics studies; more efforts have been done to develop techniques that allow studying nuclear structure. Chromosome conformation capture (3C) has been set up in 2002 from Dekker and from that moment great investments were made to develop genomics variants of 3C technology (4C, 5C, Hi-C) providing new tools to investigate the shape of the genome in a more systematic and unbiased manner. 3C method allows scientists to fix dynamic and variable 3D interactions in nuclear space, and consequently to study which sequences interact, how a gene is regulated by different and distant enhancer, or how a set of enhancer could regulate transcriptional units; to follow the conformation that mediates regulation change in development; and to evaluate if this fine epigenetic mechanism is impaired in disease condition
Going Beyond Counting First Authors in Author Co-citation Analysis
No full textThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
How Polycomb-Mediated Cell Memory Deals With a Changing Environment
No full textCells and tissues are continuously exposed to a changing microenvironment, hence the necessity of a flexible modulation of gene expression that in complex organism have been achieved through specialized chromatin mechanisms. Chromatin-based cell memory enables cells to maintain their identity by fixing lineage specific transcriptional programs, ensuring their faithful transmission through cell division; in particular PcG-based memory system evolved to maintain the silenced state of developmental and cell cycle genes. In evolution the complexity of this system have increased, particularly in vertebrates, indicating combinatorial and dynamic properties of Polycomb proteins, in some cases even overflowing outside the cell nucleus. Therefore, their function may not be limited to the imposition of rigid states of genetic programs, but on the ability to recognize signals and allow plastic transcriptional changes in response to different stimuli. Here, we discuss the most novel PcG mediated memory functions in facing and responding to the challenges posed by a fluctuating environment
Dispelling the Myths Behind First-author Citation Counts
No full textWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
An Algorithm for the Analysis of the 3D Spatial Organization of the Genome.
No full textWe present an algorithm, and its MATLAB implementation, based on mathematical methods to detect and localize 3D multicolor DNA FISH spots in fluorescence cell image z-stacks. This algorithm provides a method to measure the relative positioning of spots in the nucleus and inter-spot distances with the aim to enrich our understanding of the 3D spatial organization of the genome within the cell nucleus
3D multicolor DNA fish tool to study nuclear architecture in human primary cells
No full textA major question in cell biology is genomic organization within the nuclear space and how chromatin architecture can influence processes such as gene expression, cell identity and differentiation. Many approaches developed to study the 3D architecture of the genome can be divided into two complementary categories: chromosome conformation capture based technologies (C-technologies) and imaging. While the former is based on capturing the chromosome conformation and proximal DNA interactions in a population of fixed cells, the latter, based on DNA fluorescence in situ hybridization (FISH) on 3D-preserved nuclei, allows contemporary visualization of multiple loci at a single cell level (multicolor), examining their interactions and distribution within the nucleus (3D multicolor DNA FISH). The technique of 3D multicolor DNA FISH has a limitation of visualizing only a few predetermined loci, not permitting a comprehensive analysis of the nuclear architecture. However, given the robustness of its results, 3D multicolor DNA FISH in combination with 3D-microscopy and image reconstruction is a possible method to validate C-technology based results and to unambiguously study the position and organization of specific loci at a single cell level. Here, we propose a step by step method of 3D multicolor DNA FISH suitable for a wide range of human primary cells and discuss all the practical actions, crucial steps, notions of 3D imaging and data analysis needed to obtained a successful and informative 3D multicolor DNA FISH within different biological contexts
Mutations in the coding region of the FOXL2 gene are not a major cause of idiopathic premature ovarian failure
No full textPremature ovarian failure (POF) is a heterogeneous disorder whose aetiology is
still unknown. Recently, the autosomal FOXL2 gene, highly expressed in the adult
ovary, has been correlated with the disorder. FOXL2 mutations, causing a
truncation of the FOXL2 protein in the forkhead domain or in the poly-Ala tract
lead to blepharophimosis-ptosis-epicanthus-inversus syndrome associated with POF
(BPES I). Interestingly, in two out of 70 idiopathic POF patients, a 30 bp
deletion (898-927del) and a missense mutation (1009T-->A) were identified. To
further evaluate the correlation between POF and FOXL2 mutations, 120
phenotypically normal women affected by POF were analysed by direct sequencing
of the FOXL2 coding region. The analysis did not reveal any mutation in the 240
analysed chromosomes, indicating that mutations in the FOXL2 coding region are
rarely associated with non-syndromic PO
Mutations in the coding region of the FOXL2 gene are not a major cause of idiopathic premature ovarian failure
No full textEngaging active stakeholders in the social enterprise: Evidence of social values as a challenge to organizational identity.
No full textThe pressure on firms to be socially responsible continuously increases. Social
initiatives, however, are not without controversy. Drawing from stakeholder theory, and
the main literature on engagement and social enterprise, our research aims to verify the
role of the social dimension as a driver of active stakeholder engagement in the social
enterprise domain. The research project is based on a survey involving 268 active
stakeholders (both internal and external) of 12 Italian SEs. Based on the structural
equation modelling technique, our results provide empirical evidence on the
antecedents of engagement, distinguishing between job and organization engagement.
They confirm the relevance of the social meaningfulness of the work and of adherence to
the social values of the organization as pertinent and appropriate engagement drivers
- …
