Chromosome condensation induced by geminivirus infection of mature plant cells

2000 ◽  
Vol 113 (7) ◽  
pp. 1149-1160 ◽  
Author(s):  
H.W. Bass ◽  
S. Nagar ◽  
L. Hanley-Bowdoin ◽  
D. Robertson
Keyword(s):  
Coat Protein ◽  
Chromatin Condensation ◽  
Nuclear Periphery ◽  
Nuclear Architecture ◽  
Plant Cells ◽  
Image Data ◽  
Chromosome Condensation ◽  
Fish Analysis ◽  
Viral Dna ◽  
Plant Dna

Tomato golden mosaic virus (TGMV) is a geminivirus that replicates its single-stranded DNA genome through double-stranded DNA intermediates in nuclei of differentiated plant cells using host replication machinery. We analyzed the distribution of viral and plant DNA in nuclei of infected leaves using fluorescence in situ hybridization (FISH). TGMV-infected nuclei showed up to a sixfold increase in total volume and displayed a variety of viral DNA accumulation patterns. The most striking viral DNA patterns were bright, discrete intranuclear compartments, but diffuse nuclear localization was also observed. Quantitative and spatial measurements of high resolution 3-dimensional image data revealed that these compartments accounted for 1–18% of the total nuclear volume or 2–45% of the total nuclear FISH signals. In contrast, plant DNA was concentrated around the nuclear periphery. In a significant number of nuclei, the peripheral chromatin was organized as condensed prophase-like fibers. A combination of FISH analysis and indirect immunofluorescence with viral coat protein antibodies revealed that TGMV virions are associated with the viral DNA compartments. However, the coat protein antibodies failed to cross react with some large viral DNA inclusions, suggesting that encapsidation may occur after significant viral DNA accumulation. Infection by a TGMV mutant with a defective coat protein open reading frame resulted in fewer and smaller viral DNA-containing compartments. Nevertheless, nuclei infected with the mutant virus increased in size and in some cases showed chromosome condensation. Together, these results established that geminivirus infection alters nuclear architecture and can induce plant chromatin condensation characteristic of cells arrested in early mitosis.

scholarly journals Nuclear Position and Shape Deformation of Chromosome 8 Territories in Pancreatic Ductal Adenocarcinoma

2011 ◽  
Vol 34 (1-2) ◽  
pp. 21-33 ◽  
Author(s):  
Sylvia Timme ◽  
Eberhard Schmitt ◽  
Stefan Stein ◽  
Jutta Schwarz-Finsterle ◽  
Jenny Wagner ◽  
...  
Keyword(s):  
Nuclear Periphery ◽  
Nuclear Architecture ◽  
Neoplastic Cell ◽  
Carcinoma Cells ◽  
Chromosome 8 ◽  
Ductal Adenocarcinoma ◽  
Cell Nuclei ◽  
Formalin Fixed Paraffin ◽  
Transcriptional Alterations ◽  
Functional Relevance

Cell type specific radial positioning of chromosome territories (CTs) and their sub-domains in the interphase seem to have functional relevance in non-neoplastic human nuclei, while much less is known about nuclear architecture in carcinoma cells and its development during tumor progression. We analyzed the 3D-architecture of the chromosome 8 territory (CT8) in carcinoma and corresponding non-neoplastic ductal pancreatic epithelium. Fluorescence-in-situ-hybridization (FISH) with whole chromosome painting (WCP) probes on sections from formalin-fixed, paraffin wax-embedded tissues from six patients with ductal adenocarcinoma of the pancreas was used. Radial positions and shape parameters of CT8 were analyzed by 3D-microscopy. None of the parameters showed significant inter-individual changes. CT8 was localized in the nuclear periphery in carcinoma cells and normal ductal epithelial cells. Normalized volume and surface of CT8 did not differ significantly. In contrast, the normalized roundness was significantly lower in carcinoma cells, implying an elongation of neoplastic cell nuclei. Unexpectedly, radial positioning of CT8, a dominant parameter of nuclear architecture, did not change significantly when comparing neoplastic with non-neoplastic cells. A significant deformation of CT8, however, accompanies nuclear atypia of carcinoma cells. This decreased roundness of CTs may reflect the genomic and transcriptional alterations in carcinoma.

Nuclear Dynamics and Gene Activation during Erythroid Maturation.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 819-819
Author(s):  
M. A. Bender ◽  
Tobias Ragoczy ◽  
Rachel Byron ◽  
Agnes Telling ◽  
Mark Groudine
Keyword(s):  
Rna Polymerase Ii ◽  
Fetal Liver ◽  
Nuclear Periphery ◽  
Gene Activation ◽  
Erythroid Differentiation ◽  
Mouse Strains ◽  
Fish Analysis ◽  
Primary Transcript ◽  
Nuclear Dynamics ◽  
Erythroid Maturation

Abstract We have investigated the relationships among nuclear positioning, association with RNA polymerase II (PolII) and expression of the murine β-globin locus during erythroid differentiation, as well as the role of the locus control region (LCR) in these processes. Fetal liver cells from wildtype and LCR-deletion mouse strains were stained with a panel of antibodies, and flow cytometry was used to define and isolate cells from four stages of erythropoiesis spanning pro-erythroblasts (stage 1) to orthochromatic normoblasts and nucleated RBC (stage 4). DNA FISH analyses reveal that with increasing erythroid maturation the β-globin locus is less likely to be located in the nuclear periphery. Immuno-FISH demonstrates that PolII speckles initially are spread diffusely throughout the nucleus, with the exception of extreme periphery. With erythroid maturation the number of speckles decline and become more centrally located. Combined DNA FISH /PolII immuno-staining reveals an increase in co-localization of PolII and β-globin alleles during erythroid maturation. These results are consistent with a model in which the locus is more likely to be in the nuclear periphery and away from PolII foci prior to activation, and with maturation, the locus re-locates more centrally and associates with PolII speckles, leading to β-globin activation. To investigate this model, α and β-globin primary transcript FISH were performed and revealed that while β activation lags behind α, both start at stage 2 and increase with maturation. Notably, activation occurs prior to the increased localization of the locus away from the periphery during maturation, suggesting that localization of the locus away from the periphery may be secondary to the redistribution of PolII during erythroid maturation. Analysis of sorted erythroid cells from mice with a targeted deletion of the LCR reveals less co-localization of the β-globin locus and PolII speckles and loss of re-localization away from the periphery during maturation. Moreover, primary transcript FISH analysis of sorted cells from mice lacking the LCR and those carrying a combined deletion of HS 2 and 3 suggests that the LCR affects the likelihood that an allele is expressed, as well as the amount of transcript generated during periods of expression. Taken together our results suggest that the LCR plays a role in locating to, or stabilization of, interactions between the locus and PolII speckles, increasing the probability of β-globin expression, after which the LCR also affects the rate of transcription.

scholarly journals Viral DNA Sensors IFI16 and Cyclic GMP-AMP Synthase Possess Distinct Functions in Regulating Viral Gene Expression, Immune Defenses, and Apoptotic Responses during Herpesvirus Infection

mBio ◽  
2016 ◽  
Vol 7 (6) ◽  
Author(s):  
Benjamin A. Diner ◽  
Krystal K. Lum ◽  
Jared E. Toettcher ◽  
Ileana M. Cristea
Keyword(s):  
Gene Expression ◽  
Live Cell Imaging ◽  
Nuclear Periphery ◽  
Viral Gene Expression ◽  
Live Cell ◽  
Viral Gene ◽  
Viral Dna ◽  
Herpes Simplex Virus 1 ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACTThe human interferon-inducible protein IFI16 is an important antiviral factor that binds nuclear viral DNA and promotes antiviral responses. Here, we define IFI16 dynamics in space and time and its distinct functions from the DNA sensor cyclic dinucleotide GMP-AMP synthase (cGAS). Live-cell imaging reveals a multiphasic IFI16 redistribution, first to viral entry sites at the nuclear periphery and then to nucleoplasmic puncta upon herpes simplex virus 1 (HSV-1) and human cytomegalovirus (HCMV) infections. Optogenetics and live-cell microscopy establish the IFI16 pyrin domain as required for nuclear periphery localization and oligomerization. Furthermore, using proteomics, we define the signature protein interactions of the IFI16 pyrin and HIN200 domains and demonstrate the necessity of pyrin for IFI16 interactions with antiviral proteins PML and cGAS. We probe signaling pathways engaged by IFI16, cGAS, and PML using clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9-mediated knockouts in primary fibroblasts. While IFI16 induces cytokines, only cGAS activates STING/TBK-1/IRF3 and apoptotic responses upon HSV-1 and HCMV infections. cGAS-dependent apoptosis upon DNA stimulation requires both the enzymatic production of cyclic dinucleotides and STING. We show that IFI16, not cGAS or PML, represses HSV-1 gene expression, reducing virus titers. This indicates that regulation of viral gene expression may function as a greater barrier to viral replication than the induction of antiviral cytokines. Altogether, our findings establish coordinated and distinct antiviral functions for IFI16 and cGAS against herpesviruses.IMPORTANCEHow mammalian cells detect and respond to DNA viruses that replicate in the nucleus is poorly understood. Here, we decipher the distinct functions of two viral DNA sensors, IFI16 and cGAS, during active immune signaling upon infection with two herpesviruses, herpes simplex virus 1 (HSV-1) and human cytomegalovirus (HCMV). We show that IFI16 rapidly oligomerizes at incoming herpesvirus genomes at the nuclear periphery to transcriptionally repress viral gene expression and limit viral replicative capacity. We further demonstrate that IFI16 does not initiate upstream activation of the canonical STING/TBK-1/IRF3 signaling pathway but is required for downstream antiviral cytokine expression. In contrast, we find that, upon DNA sensing during herpesvirus infection, cGAS triggers apoptosis in a STING-dependent manner. Our live-cell imaging, mass spectrometry-based proteomics, CRISPR-based cellular assays, and optogenetics underscore the value of integrative approaches to uncover complex cellular responses against pathogens.

scholarly journals Chromatin condensation during apoptosis requires ATP

1996 ◽  
Vol 318 (3) ◽  
pp. 749-752 ◽  
Author(s):  
George E. N. KASS ◽  
John E. ERIKSSON ◽  
Marianne WEIS ◽  
Sten ORRENIUS ◽  
Sek C. CHOW
Keyword(s):  
Morphological Changes ◽  
Chromatin Condensation ◽  
Nuclear Periphery ◽  
High Molecular Mass ◽  
Jurkat Cells ◽  
Apoptotic Body ◽  
In Vitro System ◽  
Body Formation ◽  
Isolated Rat

The processes leading to morphological changes of the chromatin in cells that undergo apoptosis are presently unclear. We have recently shown that chromatin fragmentation and the nuclear morphological changes typically seen in apoptosis were reproduced in an in vitro system comprised of isolated rat thymocyte nuclei incubated in the presence of a lysate from Fas/APO-1-stimulated JURKAT cells [Chow, Weis, Kass, Holmström, Eriksson and Orrenius (1995) FEBS Lett. 364, 134–138]. Using this in vitro system, we now report that the presence of ATP is necessary for chromatin condensation, its movement to the nuclear periphery and apoptotic body formation. In clear contrast, chromatin cleavage into high-molecular-mass and oligonucleosomal-length DNA fragments induced by lysates derived from Fas/APO-1-activated JURKAT cells did not require the presence of ATP. The induction of these morphological changes by ATP could not be substituted by the analogues, adenosine 5´-[β,γ-methylene]triphosphate and adenosine 5´-[α,β-methylene]-triphosphate, AMP, cAMP and UTP. However, adenosine 5´-[γ-thio]triphosphate, and to a lesser degree GTP and ADP, could partially replace ATP in inducing nuclear apoptotic morphological changes. It is concluded that ATP is essential for the morphological changes occurring in nuclei during apoptosis, but not for DNA fragmentation.

scholarly journals Quiescent Saccharomyces cerevisiae forms telomere hyperclusters at the nuclear membrane vicinity through a multifaceted mechanism involving Esc1, the Sir complex, and chromatin condensation

2016 ◽  
Vol 27 (12) ◽  
pp. 1875-1884 ◽  
Author(s):  
Damien Laporte ◽  
Fabien Courtout ◽  
Sylvain Tollis ◽  
Isabelle Sagot
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nuclear Membrane ◽  
Chromatin Condensation ◽  
Histone H1 ◽  
Chromosome Condensation ◽  
Histone H4 ◽  
Microtubule Bundle ◽  
Quiescent Cells ◽  
Nuclear Reorganization ◽  
Sir Complex

Like other eukaryotes, Saccharomyces cerevisiae spatially organizes its chromosomes within the nucleus. In G1 phase, the yeast’s 32 telomeres are clustered into 6–10 foci that dynamically interact with the nuclear membrane. Here we show that, when cells leave the division cycle and enter quiescence, telomeres gather into two to three hyperclusters at the nuclear membrane vicinity. This localization depends on Esc1 but not on the Ku proteins. Telomere hypercluster formation requires the Sir complex but is independent of the nuclear microtubule bundle that specifically assembles in quiescent cells. Importantly, mutants deleted for the linker histone H1 Hho1 or defective in condensin activity or affected for histone H4 Lys-16 deacetylation are impaired, at least in part, for telomere hypercluster formation in quiescence, suggesting that this process involves chromosome condensation. Finally, we establish that telomere hypercluster formation is not necessary for quiescence establishment, maintenance, and exit, raising the question of the physiological raison d’être of this nuclear reorganization.

scholarly journals Heterochromatin rewiring and domain disruption-mediated chromatin compaction during erythropoiesis

2021 ◽  
Author(s):  
Dong Li ◽  
Fan Wu ◽  
Shuo Zhou ◽  
Xiaojun Huang ◽  
Hsiang-Ying Sherry Lee
Keyword(s):  
Molecular Mechanisms ◽  
Chromatin Condensation ◽  
Nuclear Periphery ◽  
Chromatin Modification ◽  
Three Dimensional ◽  
Molecular Characteristics ◽  
Structural Factors ◽  
Erythroid Progenitors ◽  
Chromatin Compaction ◽  
Active Transcription

Development of mammalian red blood cells involves progressive chromatin compaction and subsequent enucleation in terminal stages of differentiation, but the molecular mechanisms underlying the three-dimensional chromatin reorganization and compaction remains obscure. Here, we systematically analyze the distinct features of higher-order chromatin in purified populations of primary human erythroblasts. Our results reveal that while heterochromatin regions undergo substantial compression, select transcription competent regions with active transcription signature are preferentially maintained to achieve a highly-compacted yet functional chromatin state in terminal erythropoiesis, which is about 20-30% of the nuclear volume compared to that of erythroid progenitors. While the partition of euchromatic and heterochromatic regions (compartment A and B) remain mostly unchanged, H3K9me3 marks relocalize to the nuclear periphery and a significant number of H3K9me3 long-range interactions are formed in the three-dimensional rewiring during terminal erythroid chromatin condensation. Moreover, ~63% of the topologically associating domain (TAD) boundaries are disrupted, while certain TADs with active chromatin modification are selectively maintained during terminal erythropoiesis. The most well-maintained TADs are enriched for chromatin structural factors CTCF and SMC3, as well as factors and marks of the active transcription state. Finally, we demonstrate that the erythroid master regulator GATA1 involves in safeguarding select essential chromatin domains during terminal erythropoiesis. Our study therefore delineate the molecular characteristics of a development-driven chromatin compaction process, which reveals transcription competence as a key determinant of the select domain maintenance to ensure appropriate gene expression during immense chromatin compaction.

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