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Reorganization of chromosome architecture in replicative cellular senescence

2016-02-05 14:02:23

Science Advances; 5 February 2016: DOI:10.1126/sciadv.1500882

Steven W. Criscione, Marco De Cecco, Benjamin Siranosian, Yue Zhang, Jill A. Kreiling, John M. Sedivy, and Nicola Neretti



Abstract


Replicative cellular senescence is a fundamental biological process characterized by an irreversible arrest of proliferation. Senescent cells accumulate a variety of epigenetic changes, but the three-dimensional (3D) organization of their chromatin is not known. We applied a combination of whole-genome chromosome conformation capture (Hi-C), fluorescence in situ hybridization, and in silico modeling methods to characterize the 3D architecture of interphase chromosomes in proliferating, quiescent, and senescent cells. Although the overall organization of the chromatin into active (A) and repressive (B) compartments and topologically associated domains (TADs) is conserved between the three conditions, a subset of TADs switches between compartments. On a global level, the Hi-C interaction matrices of senescent cells are characterized by a relative loss of long-range and gain of short-range interactions within chromosomes. Direct measurements of distances between genetic loci, chromosome volumes, and chromatin accessibility suggest that the Hi-C interaction changes are caused by a significant reduction of the volumes occupied by individual chromosome arms. In contrast, centromeres oppose this overall compaction trend and increase in volume. The structural model arising from our study provides a unique high-resolution view of the complex chromosomal architecture in senescent cells.



Introduction


Replicative cellular senescence is recognized as a fundamentally important biological process with roles in aging, tumor suppression, embryonic development, tissue repair, and wound healing. In many of these contexts, cellular senescence is believed to be triggered by genotoxic stresses, leading to irreparable DNA damage (for example, telomere dysfunction arising from replicative exhaustion, or replication fork collapse elicited by activation of oncogenes). The accumulation of senescent cells in tissues has long been thought to contribute to age-related pathologies and functional decline, and recent evidence demonstrating beneficial effects of the removal of senescent cells supports this hypothesis.



The partitioning of chromosomes into accessible euchromatin and compact heterochromatin is central to the organization of the genome, and cellular senescence has been associated with numerous alterations to the chromatin landscape. Compacted chromatin domains, known as senescence-associated heterochromatin foci (SAHF), are formed in some but not all senescent cell nuclei. DNA in heterochromatic regions becomes globally hypomethylated, whereas some CpG islands become hypermethylated. Expression of lamin B1 decreases in senescent cells and has been associated with a loss of peripheral heterochromatin, whereas centromeric regions become dramatically distended. Using formaldehyde-assisted isolation of regulatory elements (FAIRE), we found decreased chromatin accessibility in euchromatic regions, including gene promoters and enhancers, whereas heterochromatic regions became more accessible. It has been proposed that some of these chromatin changes contribute to the gene expression patterns characteristic of senescent cells and reinforce the irreversibility of the arrest.



These dramatic chromatin alterations are very likely to affect nuclear architecture, but the three-dimensional (3D) organization of chromosomes in senescent cells is not known. Recently developed high-throughput chromosome conformation capture methods, such as Hi-C and 5C, allow a high-resolution interrogation of 3D chromosome structure. Hi-C experiments on interphase cells revealed a fundamental, large-scale organization of the genome into two types of compartments, designated as A and B. These compartments are approximately 5 Mb in size and appear to be cell type–specific. Each chromosome is composed of multiple A and B compartments, which preferentially interact in cis (A with A, and B with B). The A compartments correlate with early replicating, euchromatic regions, whereas the B compartments correlate with heterochromatin. At higher resolution (<2 Mb), the genome is organized into topologically associated domains (TADs), which can be identified by the frequent interactions between loci located within these regions (26–28). TADs can be located in either A or B compartments, and the activity of the genes in a TAD typically reflects its placement (that is, active in A and repressed in B). However, unlike compartments, TADs are highly conserved across cell types and even species, and are believed to constitute a fundamental organizational unit of the genome. Supporting their organizational role, TAD boundaries are highly correlated with replication timing domains.



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Key Words


Gene regulation | Genetic variation | Reporter genes | Transcriptional regulatory elements