Dataset Information

Interplay of lamin A and lamin B LADs on the radial positioning of chromatin.

ABSTRACT: Immunosuppressive drugs such as cyclosporin A (CsA) can elicit hepatotoxicity by affecting gene expression. Here, we address the link between CsA and large-scale chromatin organization in HepG2 hepatocarcinoma cells. We show the existence of lamina-associated domains (LADs) interacting with lamin A, lamin B, or both. These 'A-B', 'A-only' and 'B-only' LADs display distinct fates after CsA treatment: A-B LADs remain constitutive or lose A, A-only LADs mainly lose A or switch to B, and B-only LADs remain B-only or acquire A. LAD rearrangement is overall uncoupled from changes in gene expression. Three-dimensional (3D) genome modeling predicts changes in radial positioning of LADs as LADs switch identities, which are corroborated by fluorescence in situ hybridization. Our results reveal interplay between A- and B-type lamins on radial locus positioning, suggesting complementary contributions to large-scale genome architecture. The data also unveil a hitherto unsuspected impact of cytotoxic drugs on genome conformation.Abbreviations: ChIP-seq: chromatin immunoprecipitation sequencing; CsA: cyclosporin A; FISH; fluorescence in situ hybridization; ICMT: isoprenylcysteine methyltransferase; LAD: lamina-associated domain; TAD: topologically-associated domain.

SUBMITTER: Forsberg F

PROVIDER: S-EPMC6363278 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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Interplay of lamin A and lamin B LADs on the radial positioning of chromatin.

Forsberg Frida F Brunet Annaël A Ali Tharvesh M Liyakat TML Collas Philippe P

Nucleus (Austin, Tex.) 20191201 1

Immunosuppressive drugs such as cyclosporin A (CsA) can elicit hepatotoxicity by affecting gene expression. Here, we address the link between CsA and large-scale chromatin organization in HepG2 hepatocarcinoma cells. We show the existence of lamina-associated domains (LADs) interacting with lamin A, lamin B, or both. These 'A-B', 'A-only' and 'B-only' LADs display distinct fates after CsA treatment: A-B LADs remain constitutive or lose A, A-only LADs mainly lose A or switch to B, and B-only LADs ...[more]

PMID: 30663495

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Project description:G- and R-bands of metaphase chromosomes are characterized by profound differences in gene density, CG content, replication timing, and chromatin compaction. The preferential localization of gene-dense, transcriptionally active, and early replicating chromatin in the nuclear interior and of gene-poor, later replicating chromatin at the nuclear envelope has been demonstrated to be evolutionary-conserved in various cell types. Yet, the impact of different local chromatin features on the radial nuclear arrangement of chromatin is still not well understood. In particular, it is not known whether radial chromatin positioning is preferentially shaped by local gene density per se or by other related parameters such as replication timing or transcriptional activity. The interdependence of these distinct chromatin features on the linear deoxyribonucleic acid (DNA) sequence precludes a simple dissection of these parameters with respect to their importance for the reorganization of the linear DNA organization into the distinct radial chromatin arrangements observed in the nuclear space. To analyze this problem, we generated probe sets of pooled bacterial artificial chromosome (BAC) clones from HSA 11, 12, 18, and 19 representing R/G-band-assigned chromatin, segments with different gene density and gene loci with different expression levels. Using multicolor 3D flourescent in situ hybridization (FISH) and 3D image analysis, we determined their localization in the nucleus and their positions within or outside the corresponding chromosome territory (CT). For each BAC data on local gene density within 2- and 10-Mb windows, as well as GC (guanine and cytosine) content, replication timing and expression levels were determined. A correlation analysis of these parameters with nuclear positioning revealed regional gene density as the decisive parameter determining the radial positioning of chromatin in the nucleus in contrast to band assignment, replication timing, and transcriptional activity. We demonstrate a polarized distribution of gene-dense vs gene-poor chromatin within CTs with respect to the nuclear border. Whereas we confirm previous reports that a particular gene-dense and transcriptionally highly active region of about 2 Mb on 11p15.5 often loops out from the territory surface, gene-dense and highly expressed sequences were not generally found preferentially at the CT surface as previously suggested.

Project description:Purpose: to qualify and quantify the rearrangement of interactions of nuclear lamins A/C and B with the genome, in relation to nuclearradial repositioning of loci and changes in gene expression, in HepG2 cells exposed to cyclosporin A. .To compare HepG2 cell line transcriptome profiling (RNA-seq) with radially positioning of the chromatin anchored at the nuclear periphery before and after CsA treatment. Methods: To this end, we mapped lamin A and lamin B lamina-associated domains (LADs) by chromatin immunoprecipitation-sequencing (ChIP-seq) of lamin A/C (antibody sc7292x, Santa Cruz Biotechnology) and lamin B1 (antibody ab16048, Abcam), respectively. HepG2 cells were either cultured under proliferative conditions (controls) or exposed for 3 days to 10 uM cyclosporin A. We also assesed the transcriptome by RNA-seq under each condition in duplicate cultures. Results: CsA elicits accumulation of pre-lamin A in HepG2 cells, raising the hypothesis of a global rearrangement of lamin-chromatin interactions. We show here the existence of overlapping and distinct lamin A and B lamina-associated domains (which we refer to as A- and B-LADs, respectively). Whereas B-LADs largely remain constitutive, CsA elicits the formation of facultative A-LADs. Re-localization of A-LADs occur mainly on preexisting B-LADs. However, LAD rearrangement does not correlate with systematic changes in gene expression within LADs. We find a reorganization of A- and B-LADs after CsA treatment, entailing distinct contributions of A- and B- type lamins. to genome organization. Indeed, classification of lamin A LADs and lamin B LADs into properties of ‘A-only’, ‘B-only’ and ‘A-B’ LADs show an overall loss of A-only LADs after CsA treatment. A- only and B-only LADs are highly dynamic with A-B LADs being more stable.The rearrangement of lamin association after CsA treatment correlates with repositioning of loci in the nuclear space with, notably, a loss of lamin B associated with a re-localization of loci towards the nuclear center and conversely, a gain of lamin B linked to a repositioning of loci towards the nuclear periphery. Predictions on radial repositioning of LADs, and of loci within, from 3D structural models of the genome were supported by fluorescence in situ hybridization (FISH) analyses. Conclusions: We show that, while they amply co-localize on well-conserved LADs, lamins A and B also interact with distinct genomic regions that can interchange (switch) between lamin A and B upon CsA treatment. We unveil distinct lamin A and B interactions with chromatin, suggesting complementary contributions to genome conformation. Our data suggest that and lamins A and B may differentially modulate genome organization.

Dataset Information

Interplay of lamin A and lamin B LADs on the radial positioning of chromatin.

Publications

Interplay of lamin A and lamin B LADs on the radial positioning of chromatin.

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