bims-nutges 2021-08-08 papers

Issue of 2021–08–08
three papers selected by
Kanwal Tariq, Stockholm University

Cancers (Basel). 2021 Aug 02. pii: 3889. [Epub ahead of print]13(15):

Chromosomal Translocations in NK-Cell Lymphomas Originate from Inter-Chromosomal Contacts of Active rDNA Clusters Possessing Hot Spots of DSBs.

Nickolai A Tchurikov, Leonid A Uroshlev, Elena S Klushevskaya, Ildar R Alembekov, Maria A Lagarkova, Galina I Kravatskaya, Vsevolod Y Makeev, Yuri V Kravatsky.

Endogenous hot spots of DNA double-strand breaks (DSBs) are tightly linked with transcription patterns and cancer. There are nine hot spots of DSBs (denoted Pleiades) in human rDNA units that are located exclusively inside the intergenic spacer (IGS). Profiles of Pleiades coincide with the profiles of γ-H2AX, suggesting a high level of in vivo breakage inside rDNA genes. The data were confirmed by microscopic observation of the largest γ-H2AX foci inside nucleoli in interphase chromosomes. Circular chromosome conformation capture (4C) data indicate that the rDNA units often make contact with a specific set of chromosomal regions containing genes that are involved in differentiation and cancer. Interestingly, these regions also often possess hot spots of DSBs that provide the potential for Robertsonian and oncogenic translocations. In this study, we searched for translocations in which rDNA clusters are involved. The whole genome sequence (WGS) data of normal T cells and NK-cell lymphomas from the same individuals revealed numerous translocations in which Pleiades were involved. The sites of these translocations in normal T cells and in the lymphomas were mostly different, although there were also some common sites. The genes at translocations in normal cells and in lymphomas are associated with predominantly non-overlapping lists of genes that are depleted with silenced genes. Our data indicate that rDNA-mediated translocations occur at about the same frequency in the normal T cells and NK-lymphoma cells but differ at particular sites that correspond to open chromatin. We conclude that oncogenic translocations lead to dysregulation of a specific set of genes controlling development. In normal T cells and in NK cells, there are hot spots of translocations at sites possessing strong H3K27ac marks. The data indicate that Pleiades are involved in rDNA-mediated translocation.

Keywords: 4C; NK-cell lymphomas; T cells; inter-chromosomal contacts; rDNA genes; translocations

DOI: https://doi.org/10.3390/cancers13153889

Genes (Basel). 2021 Jul 16. pii: 1081. [Epub ahead of print]12(7):

Nuclear Organization during Hepatogenesis in Zebrafish Requires Uhrf1.

Bhavani P Madakashira, Chi Zhang, Filippo Macchi, Elena Magnani, Kirsten C Sadler.

Acquisition of cellular fate during development is initiated and maintained by well-coordinated patterns of gene expression that are dictated by the epigenetic landscape and genome organization in the nucleus. While the epigenetic marks that mediate developmental gene expression patterns during organogenesis have been well studied, less is known about how epigenetic marks influence nuclear organization during development. This study examines the relationship between nuclear structure, chromatin accessibility, DNA methylation, and gene expression during hepatic outgrowth in zebrafish larvae. We investigate the relationship between these features using mutants that lack DNA methylation. Hepatocyte nuclear morphology was established coincident with hepatocyte differentiation at 80 h post-fertilization (hpf), and nuclear shape and size continued to change until the conclusion of outgrowth and morphogenesis at 120 hpf. Integrating ATAC-Seq analysis with DNA methylation profiling of zebrafish livers at 120 hpf showed that closed and highly methylated chromatin occupies most transposable elements and that open chromatin correlated with gene expression. DNA hypomethylation, due to mutation of genes encoding ubiquitin-like, containing PHD and RING Finger Domains 1 (uhrf1) and DNA methyltransferase (dnmt1), did not block hepatocyte differentiation, but had dramatic effects on nuclear organization. Hepatocytes in uhrf1 mutants have large, deformed nuclei with multiple nucleoli, downregulation of nucleolar genes, and a complete lack of the nuclear lamina. Loss of lamin B2 staining was phenocopied by dnmt1 mutation. Together, these data show that hepatocyte nuclear morphogenesis coincides with organ morphogenesis and outgrowth, and that DNA methylation directs chromatin organization, and, in turn, hepatocyte nuclear shape and size during liver development.

Keywords: DNA methylation; liver development; nuclear organization; nuclear structure; organogenesis; uhrf1; zebrafish

DOI: https://doi.org/10.3390/genes12071081

J Cell Sci. 2021 Aug 01. pii: jcs258374. [Epub ahead of print]134(15):

Super-resolution microscopy of chromatin fibers and quantitative DNA methylation analysis of DNA fiber preparations.

Michal Franek, Agata Kilar, Petr Fojtík, Marie Olšinová, Aleš Benda, Vladimír Rotrekl, Martina Dvořáčková, Jíří Fajkus.

Analysis of histone variants and epigenetic marks is dominated by genome-wide approaches in the form of chromatin immunoprecipitation-sequencing (ChIP-seq) and related methods. Although uncontested in their value for single-copy genes, mapping the chromatin of DNA repeats is problematic for biochemical techniques that involve averaging of cell populations or analysis of clusters of tandem repeats in a single-cell analysis. Extending chromatin and DNA fibers allows us to study the epigenetics of individual repeats in their specific chromosomal context, and thus constitutes an important tool for gaining a complete understanding of the epigenetic organization of genomes. We report that using an optimized fiber extension protocol is essential in order to obtain more reproducible data and to minimize the clustering of fibers. We also demonstrate that the use of super-resolution microscopy is important for reliable evaluation of the distribution of histone modifications on individual fibers. Furthermore, we introduce a custom script for the analysis of methylation levels on DNA fibers and apply it to map the methylation of telomeres, ribosomal genes and centromeres.

Keywords: Chromatin; DNA; Fiber; Methylation; Microscopy

DOI: https://doi.org/10.1242/jcs.258374