92 research outputs found
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
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
Repetitive elements dynamics in cell identity programming, maintenance and disease
No full textThe days of 'junk DNA' seem to be over. The rapid progress of genomics technologies has been unveiling unexpected mechanisms by which repetitive DNA and in particular transposable elements (TEs) have evolved, becoming key issues in understanding genome structure and function. Indeed, rather than 'parasites', recent findings strongly suggest that TEs may have a positive function by contributing to tissue specific transcriptional programs, in particular as enhancer-like elements and/or modules for regulation of higher order chromatin structure. Further, it appears that during development and aging genomes experience several waves of TEs activation, and this contributes to individual genome shaping during lifetime. Interestingly, TEs activity is major target of epigenomic regulation. These findings are shedding new light on the genome-phenotype relationship and set the premises to help to explain complex disease manifestation, as consequence of TEs activity deregulation.We thank Federica Marasca and Chiara Lanzuolo for helpful comments and criticisms on the manuscript. The original work of the lab is supported by EPIGEN Italian flagship program (to BB and VO) and King Abdullah University of Science and Technology (KAUST) to VO
ANALISYS OF GENE EXPRESSION PROFILE DURING MYOGENESIS EVIDENCES DIFFERENT MOLECULAR DEFECTS AT THE BASIS OF FSHD-1 AND FSHD-2 PATHOGENESIS
No full textGerm line cell-derived pluripotent stem cells (GPSCs) are derived from spermatogonial stem cells and are similar to
embryonic stem (ES) cells in that they can proliferate intensively and differentiate into a variety of cell types. Previous
studies have revealed some inherent differences in gene expression between undifferentiated mouse ES cells and
GPSCs. Our aims were to generate functional hepatocytes from mouse GPSCs in vitro and to investigate whether the
differences in gene expression may impact on the hepatocyte differentiation capacity of the GPSCs compared with ES
cells. Mouse GPSCs and ES cells were induced to differentiate into hepatocytes through embryoid body formation, with
very high efficiency. These hepatocytes were characterized at cellular, molecular, and functional levels. The GPSCderived
hepatocytes expressed hepatic markers and were metabolically active as shown by albumin and haptoglobin
secretion, urea synthesis, glycogen storage, and indocyanine green uptake. We also performed an unprecedented DNA
microarray analysis comparing different stages of hepatocyte differentiation. Gene expression profiling demonstrated a
strong similarity between GPSC and ES cells at different stages of induced hepatic differentiation. Moreover, Pearson
correlation analysis of the microarray datasets suggested that, at late hepatic differentiation stages, the in vitro-derived
cells were closer to fetal mouse primary hepatocytes than to those obtained from neonates. We have shown for the first
time that adult GPSCs can be induced to differentiate into functional hepatocytes in vitro. Moreover, our ongoing in
vivo work shows that GPSC-derived hepatocytes can colonize the liver of monocrotaline-treated, partially
hepatectomised mice. These GPSC-derived hepatocytes thus offer great potential for cell replacement therapy for a
wide variety of liver diseases
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A Ship Incised on Slate Found Near Bodega, Sonoma County, California
No full textIn the spring of 1977, the author conducted an archaeological reconnaissance near the town of Bodega, about four miles inland from Bodega Bay in Sonoma County, California. During the course of the reconnaissance, a small, broken piece of slate (Fig. 1), incised with the picture of a sailing ship (Sonoma State University Accession No. 77-3-241), was collected from the surface of archaeological site CA-Son-290, an ethnographic village of the Bodega Miwok. Since the occurrence of incised artifacts in northern California is rare, a brief discussion of the context of the find and a description of the artifact are in order
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.Beatrice Bodega and Valerio Orlando contributed equally to this work. We thank Chiara Lanzuolo and Alice Cortesi for helpful comments and criticisms on the manuscript. The work is supported by EPIGEN Italian flagship program (to BB), Istituto Nazionale di Genetica Molecolare to BB and King Abdullah University of Science and Technology (KAUST) to VO
Evolutionary history of linked D4Z4 and Beta satellite clusters at the FSHD locus (4q35)
No full textAbstractWe performed a detailed genomic investigation of the chimpanzee locus syntenic to human chromosome 4q35.2, associated to the facioscapulohumeral dystrophy. Two contigs of approximately 150kb and 200kb were derived from PTR chromosomes 4q35 and 3p12, respectively: both regions showed a very similar sequence organization, including D4Z4 and Beta satellite linked clusters. Starting from these findings, we derived a hypothetical evolutionary history of human 4q35, 10q26 and 3p12 chromosome regions focusing on the D4Z4–Beta satellite linked organization. The D4Z4 unit showed an open reading frame (DUX4) at both PTR 4q35 and 3p12 regions; furthermore some subregions of the Beta satellite unit showed a high degree of conservation between chimpanzee and humans. In conclusion, this paper provides evidence that at the 4q subtelomere the linkage between D4Z4 and Beta satellite arrays is a feature that appeared late during evolution and is conserved between chimpanzee and humans
Identification of RNA-protein interactions through in-vitro RNA pull-down assays
No full textRecent advances in next generation sequencing have revealed that majority of the human genome is transcribed into long and short RNA (ncRNA) transcripts. Many ncRNAs function by interacting with proteins and forming regulatory complexes. RNA-protein interactions are vital in controlling core cellular processes like transcription and translation. Therefore identifying proteins that interact with ncRNAs is central to deciphering ncRNA functions. Here we describe a RNA-protein pull-down assay, which enable the identification of proteins that interact with a RNA under study. As an example we describe pull-down of proteins interacting with ncRNA XIST, which assists in the recruitment of the polycomb repressive complex-2 (PRC2) and drives X-chromosomal inactivation. <br/
PD recurrence in cavities at different energizing methods
No full textElectrical Engineering, Mathematics and Computer Scienc
A Versatile Pipeline for Analyzing Dynamic Changes in Nuclear Bodies in a Variety of Cell Types
No full textVarious nuclear processes, such as transcriptional control, occur within discrete structures known as foci that are discernable through the immunofluorescence technique. Investigating the dynamics of these foci under diverse cellular conditions via microscopy yields valuable insights into the molecular mechanisms governing cellular identity and functions. However, performing immunofluorescence assays across different cell types and assessing alterations in the assembly, diffusion, and distribution of these foci present numerous challenges. These challenges encompass complexities in sample preparation, determination of parameters for analyzing imaging data, and management of substantial data volumes. Moreover, existing imaging workflows are often tailored for proficient users, thereby limiting accessibility to a broader audience. In this study, we introduce an optimized immunofluorescence protocol tailored for investigating nuclear proteins in different human primary T cell types that can be customized for any protein of interest and cell type. Furthermore, we present a method for unbiasedly quantifying protein staining, whether they form distinct foci or exhibit a diffuse nuclear distribution. Our proposed method offers a comprehensive guide, from cellular staining to analysis, leveraging a semi-automated pipeline developed in Jython and executable in Fiji. Furthermore, we provide a user-friendly Python script to streamline data management, publicly accessible on a Google Colab notebook. Our approach has demonstrated efficacy in yielding highly informative immunofluorescence analyses for proteins with diverse patterns of nuclear organization across different contexts
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