MadID, a Versatile Approach to Map Protein-DNA Interactions, Highlights Telomere-Nuclear Envelope Contact Sites in Human Cells

Sobecki, Michal; Souaid, Charbel; Boulay, Jocelyne; Guérineau, Vincent; Noordermeer, Daan; Crabbé, Laure

doi:10.1016/j.celrep.2018.11.027

Cited by 24 publications

(39 citation statements)

References 31 publications

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“…However, the extent of NE blebbing with deep membrane invaginations reaching the nucleus center made it difficult to evaluate telomere localization with regards to the nuclear lamina by microscopy. To solve this issue, we applied MadID, a proximity labeling technique that we recently developed to probe telomere-NE contact sites 35 , 42 . MadID relies on the expression of a bacterial methyltransferase called M.EcoGII, which catalyzes the methylation of adenine residues (m6A).…”

Section: Resultsmentioning

confidence: 99%

“…In interphase cells, human telomeres are organized close to the center of the nucleus 30 , 31 , but a few late-replicating telomeres are anchored to the NE 32 , 33 . In contrast, a large subset of telomeres is tethered to the NE during post-mitotic nuclear assembly 34 , in close proximity to Lamin B1 35 . Connection between telomeres and lamins also occurs in the nucleoplasm and plays a role in telomere protection 36 .…”

Section: Introductionmentioning

confidence: 99%

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Progerin impairs 3D genome organization and induces fragile telomeres by limiting the dNTP pools

Kychygina

Dall’Osto

Allen

et al. 2021

Sci Rep

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Chromatin organization within the nuclear volume is essential to regulate many aspects of its function and to safeguard its integrity. A key player in this spatial scattering of chromosomes is the nuclear envelope (NE). The NE tethers large chromatin domains through interaction with the nuclear lamina and other associated proteins. This organization is perturbed in cells from Hutchinson–Gilford progeria syndrome (HGPS), a genetic disorder characterized by premature aging features. Here, we show that HGPS-related lamina defects trigger an altered 3D telomere organization with increased contact sites between telomeres and the nuclear lamina, and an altered telomeric chromatin state. The genome-wide replication timing signature of these cells is perturbed, with a shift to earlier replication for regions that normally replicate late. As a consequence, we detected a higher density of replication forks traveling simultaneously on DNA fibers, which relies on limiting cellular dNTP pools to support processive DNA synthesis. Remarkably, increasing dNTP levels in HGPS cells rescued fragile telomeres, and improved the replicative capacity of the cells. Our work highlights a functional connection between NE dysfunction and telomere homeostasis in the context of premature aging.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Progerin impairs 3D genome organization and induces fragile telomeres by limiting the dNTP pools

Kychygina

Dall’Osto

Allen

et al. 2021

Sci Rep

Self Cite

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“…Additionally, this protocol reduces the number of steps required by using the initial ligated adapter directly for sequencing, instead of separating amplification of methylated fragments from later sequencing library preparation (as in [7]), and produces a more interpretable output of read count at each GATC instead of being smeared out into a peak. Further increasing the frequency with which binding can be detected could be achieved by combining these Dam mutants with K9A, which allows Dam to methylate at sequences other than GATC and detecting the resulting methylation by immunoprecipitation [22, 23].…”

Section: Discussionmentioning

confidence: 99%

Dam mutants provide improved sensitivity and spatial resolution for profiling transcription factor binding

Szczesnik

Sherwood

2019

Epigenetics & Chromatin

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DamID, in which a protein of interest is fused to Dam methylase, enables mapping of protein-DNA binding through readout of adenine methylation in genomic DNA. DamID offers a compelling alternative to chromatin immunoprecipitation sequencing (ChIP-Seq), particularly in cases where cell number or antibody availability is limiting. This comes at a cost, however, of high non-specific signal and a lowered spatial resolution of several kb, limiting its application to transcription factor-DNA binding. Here we show that mutations in Dam, when fused to the transcription factor Tcf7l2, greatly reduce non-specific methylation. Combined with a simplified DamID sequencing protocol, we find that these Dam mutants allow for accurate detection of transcription factor binding at a sensitivity and spatial resolution closely matching that seen in ChIP-seq. Electronic supplementary material The online version of this article (10.1186/s13072-019-0273-x) contains supplementary material, which is available to authorized users.

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“…After mitosis in somatic cells, at the stage of NE reassembly, telomeres are transiently enriched at the nuclear periphery [319]. Recently, contact between telomeres and the nuclear rim was also detected using the MadID approach in G1/S arrested HeLa cells [320]. A subset of telomeres is also found close to the nuclear periphery during replication [321].…”

Section: Lamins and Telomere Maintenancementioning

confidence: 99%

A Link between Replicative Stress, Lamin Proteins, and Inflammation

Willaume

Rass

Fontanilla-Ramirez

et al. 2021

Genes

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Double-stranded breaks (DSB), the most toxic DNA lesions, are either a consequence of cellular metabolism, programmed as in during V(D)J recombination, or induced by anti-tumoral therapies or accidental genotoxic exposure. One origin of DSB sources is replicative stress, a major source of genome instability, especially when the integrity of the replication forks is not properly guaranteed. To complete stalled replication, restarting the fork requires complex molecular mechanisms, such as protection, remodeling, and processing. Recently, a link has been made between DNA damage accumulation and inflammation. Indeed, defects in DNA repair or in replication can lead to the release of DNA fragments in the cytosol. The recognition of this self-DNA by DNA sensors leads to the production of inflammatory factors. This beneficial response activating an innate immune response and destruction of cells bearing DNA damage may be considered as a novel part of DNA damage response. However, upon accumulation of DNA damage, a chronic inflammatory cellular microenvironment may lead to inflammatory pathologies, aging, and progression of tumor cells. Progress in understanding the molecular mechanisms of DNA damage repair, replication stress, and cytosolic DNA production would allow to propose new therapeutical strategies against cancer or inflammatory diseases associated with aging. In this review, we describe the mechanisms involved in DSB repair, the replicative stress management, and its consequences. We also focus on new emerging links between key components of the nuclear envelope, the lamins, and DNA repair, management of replicative stress, and inflammation.

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MadID, a Versatile Approach to Map Protein-DNA Interactions, Highlights Telomere-Nuclear Envelope Contact Sites in Human Cells

Cited by 24 publications

References 31 publications

Progerin impairs 3D genome organization and induces fragile telomeres by limiting the dNTP pools

Progerin impairs 3D genome organization and induces fragile telomeres by limiting the dNTP pools

Dam mutants provide improved sensitivity and spatial resolution for profiling transcription factor binding

A Link between Replicative Stress, Lamin Proteins, and Inflammation

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