Biology subjects

Lleres, D.

Publications and source records attributed to Lleres, D..

5 recordsLinked to original sources

bioRxiv2023

Cell-cycle dependent inhibition of BRCA1 signaling by the lysine methyltransferase SET8

The cell-cycle regulated methyltransferase SET8 is the sole enzyme responsible for the mono-methylation of histone H4 at lysine 20 (H4K20) that is the substrate for di- and tri-methylation mainly by SUV4-20Hs enzymes. Both SET8 and SUV4-20Hs have been implicated in regulating DNA repair pathway choice through the inverse affinities of BRCA1-BARD1 and 53BP1 complexes for disparate methylation states of H4K20. However, the precise and respective functions of each H4K20 methyltransferases in DNA repair pathways remained to be clarified. Here, we show that SET8 acts as a potent chromatin inhibitor of homologous recombination and that its timely degradation during DNA replication is essential for the spontaneous nuclear focal accumulation of BRCA1 and RAD51 complexes during S-phase. Strikingly, the anti-recombinogenic function of SET8 is independent of SUV4-20H activity but requires the subsequent recruitment of the ubiquitin ligase RNF168. Moreover, we show that SET8-induced BRCA1 inhibition is not necessarily related to the loss of BARD1 binding to unmethylated histone H4K20. Instead, it is largely caused by the accumulation of 53BP1 in a manner depending on the concerted activities of SET8 and RNF168 on chromatin. Conversely, the lack of SET8 and H4K20 mono-methylation on newly assembly chromatin after DNA replication led to the untimely accumulation of BRCA1 on chromatin at the subsequent G1 phase. Altogether, these results establish the de novo activity of SET8 on chromatin as a primordial epigenetic lock of BRCA1-mediated HR pathway during the cell cycle.

molecular biology↗

bioRxiv2020

Cooperation between Caenorhabditis elegans COMPASS and condensin in germline chromatin organization

Deposition of histone H3 lysine 4 (H3K4) methylation at promoters by SET1/COMPASS is associated with context-dependent effects on gene expression and local changes in chromatin organization. Whether SET1/COMPASS also contributes to higher-order chromosome structure has not been investigated. Here, we address this question by quantitative FRET (Forster resonance energy transfer)-based fluorescence lifetime imaging microscopy (FLIM) on C. elegans germ cells expressing histones H2B-eGFP and H2B-mCherry. We find that SET1/COMPASS subunits strongly influence meiotic chromosome organization, with marked effects on the close proximity between nucleosomes. We further show that inactivation of SET-2, the C. elegans homologue of SET1, or CFP-1, the chromatin targeting subunit of COMPASS, strongly enhance chromosome organization defects and loss of fertility resulting from depletion of condensin-II. Defects in chromosome morphology resulting from conditional inactivation of topoisomerase II, another structural component of chromosomes, were also aggravated in the absence of SET-2. Combined, our in vivo findings suggest a model in which the SET1/COMPASS histone methyltransferase complex plays a role in shaping meiotic chromosome in cooperation with the non-histone proteins condensin-II and topoisomerase.

genetics↗

bioRxiv2019

The 20S proteasome activator PA28γ controls the compaction of HP1β-linked heterochromatin

PA28{gamma}, a nuclear activator of the 20S proteasome, is involved in the degradation of several proteins regulating cell growth and proliferation and in the dynamics of various nuclear bodies, but its precise cellular functions remain unclear. Here, using a quantitative FLIM-FRET based microscopy assay monitoring close proximity between nucleosomes in living human cells, we show that PA28{gamma} controls chromatin compaction. We find that its depletion induces a decompaction of pericentromeric heterochromatin, similarly to that observed upon the knockdown of HP1{beta}, a key factor in heterochromatin structure. We show that PA28{gamma} is present at HP1{beta}-containing repetitive-DNA sequences abundant in heterochromatin and importantly, that HP1{beta} on its own is unable to drive chromatin compaction without the presence of PA28{gamma}. At the molecular level, we show that this novel function of PA28{gamma} is independent of its stable interaction with the 20S proteasome, and most likely depends on its ability to maintain appropriate levels of H3K9me3 and H4K20me3, histone modifications that are both involved in heterochromatin formation. Overall, our results implicate PA28{gamma} as a key factor involved in the higher order structuration of chromatin.

cell biology↗

bioRxiv2019

CTCF controls imprinted gene activity at the mouse Dlk1-Dio3 and Igf2- H19 domains by modulating allele-specific sub-TAD structure

Mammalian genomic imprinting is essential for development and provides a unique paradigm to explore intra-cellular differences in chromatin configuration. Here, we compared chromatin structure of the two conserved imprinted domains controlled by paternal DNA methylation imprints--the Igf2-H19 and the Dlk1-Dio3 domains--and assessed the involvement of the insulator protein CTCF. At both domains, CTCF binds the maternal allele of a differentially-methylated region (DMR), in addition to multiple instances of bi-allelic CTCF binding in their surrounding TAD (Topologically Associating Domain). On the paternal chromosome, bi-allelic CTCF binding alone is sufficient to structure a first level of sub-TAD organization. Maternal-specific CTCF binding at the DMRs adds a further layer of sub-TAD organization, which essentially hijacks the existing paternal sub-TAD organisation. Genome-editing experiments at the Dlk1-Dio3 locus confirm that the maternal sub-TADs are essential during development to maintain the imprinted Dlk1 gene in an inactive state on the maternal chromosome.

molecular biology↗

bioRxiv2018

A histone H4K20 methylation-mediated chromatin compaction threshold ensures genome integrity by limiting DNA replication licensing

The decompaction and re-establishment of chromatin organization immediately after mitosis is essential for genome regulation. The mechanisms underlying chromatin structure control in daughter cells are not fully understood. Here, we show that a chromatin compaction threshold in cells exiting mitosis ensures genome integrity by limiting replication licensing in G1 phase. Upon mitotic exit, appropriate chromatin relaxation is safeguarded by SET8-dependent methylation of histone H4 on lysine 20. Thus, in the absence of either SET8 or the H4K20 residue, substantial genome-wide chromatin decompaction occurs which allows excessive loading of the Origin Recognition Complex (ORC) in the daughter cells. ORC overloading stimulates aberrant recruitment of the MCM2-7 complex that promotes single-stranded DNA formation and DNA damage. Restoring chromatin compaction restrains excess replication licensing and the loss of genome integrity. Our findings identify a cell cycle-specific mechanism whereby fine-tuned chromatin relaxation suppresses excessive detrimental replication licensing and maintains genome integrity at the cellular transition from mitosis to G1 phase.

molecular biology↗