Biology subjects

Esashi, F.

Publications and source records attributed to Esashi, F..

5 recordsLinked to original sources

bioRxiv2024

DreaM: A Computational Pipeline for Enhanced Short-Read Sequence Analysis in Repetitive Genomic Regions

Mapping short sequencing reads to repetitive genomic regions, such as centromeres, presents significant challenges, primarily due to PCR duplicates, which can be erroneously mapped multiple times within these regions. Conventional bioinformatics pipelines often overlook this issue, potentially leading to misinterpretation as signal enrichment. To address this, we developed DreaM (Deduplication of Reads for Enhanced and Accurate Mapping), a computational pipeline that prioritises the preprocessing of raw sequencing data. DreaM firstly identifies and removes PCR duplicates, which is followed by read trimming to reduce noise from multiply mapped reads. When applied to ChIP-Seq and CUT&RUN datasets targeting CENP-A, a key marker of centromeres, DreaM demonstrated improved peak detection within centromeres. Overall, DreaM provides a robust solution for enhancing the analysis of DNA-protein binding sites in repetitive genomic regions using short-read sequencing.

bioinformatics↗

bioRxiv2020

The RAD51 recombinase protects mitotic chromatin in human cells

The RAD51 recombinase plays critical roles in safeguarding genome integrity, which is fundamentally important for all living cells. While interphase functions of RAD51 in repairing broken DNA and protecting stalled replication forks are well characterised, its role in mitosis remains contentious. In this study, we show that RAD51 protects under-replicated DNA in mitotic human cells and, in this way, promotes mitotic DNA synthesis (MiDAS) and successful chromosome segregation. MiDAS was globally detectable irrespective of DNA damage and was promoted by de novo RAD51 recruitment, RAD51-mediated fork protection, and RAD51 phosphorylation by the key mitotic regulator Polo-like kinase 1. Importantly, acute inhibition of RAD51-promoted MiDAS led to mitotic DNA damage, delayed anaphase onset and induced centromere fragility, revealing a mechanism that prevents the satisfaction of the spindle assembly checkpoint when chromosomal replication remains incomplete. This study hence identifies an unexpected function of RAD51 in promoting the stability of mitotic chromatin.

molecular biology↗

bioRxiv2020

Intact nucleosomal context enables chromodomain reader MRG15 to distinguish H3K36me3 from -me2

A wealth of in vivo evidence demonstrates the physiological importance of histone H3 trimethylation at lysine 36 (H3K36me3), to which chromodomain-containing proteins, such as MRG15, bind preferentially compared to their dimethyl (H3K36me2) counterparts. However, in vitro studies using isolated H3 peptides have failed to recapitulate a causal interaction. Here, we show that MRG15 can clearly discriminate between synthetic, fully intact model nucleosomes containing H3K36me2 and H3K36me3. MRG15 docking studies, along with experimental observations and nucleosome structure analysis suggest a model where the H3K36 side chain is sequestered in intact nucleosomes via a hydrogen bonding interaction with the DNA backbone, which is abrogated when the third methyl group is added to form H3K36me3. Hence, this mechanism provides a methyl-switch for contextdependent reader selectivity. These results highlight the importance of such intra-chromatin interactions in understanding epigenetic regulation, a feature which is absent in commonly-used peptide or histone-only models.

biochemistry↗

bioRxiv2019

Lysine acetylation promotes PALB2 chromatin association to maintain genome stability

The tumour suppressor PALB2 stimulates error-free repair of DNA breaks, whilst its steady-state chromatin association protects active genes from genotoxic stress. Here, we report that the lysine acetyltransferases 2A and 2B (KAT2A/B), commonly known to promote transcriptional activation, acetylate the PALB2 chromatin association motif (ChAM), providing a dynamic regulatory mechanism for PALB2. ChAM acetylation within a cluster of seven lysine residues (7K), detected in the chromatin-enriched fraction in undamaged cells, enhanced its association with nucleosomes while decreasing its non-specific binding to naked DNA. DNA damage triggered a rapid deacetylation of ChAM and a concomitant increase in PALB2 mobility. Significantly, a 7K-null mutation, which hindered ChAM binding to both nucleosomes and DNA, conferred deficiency in DNA repair and hypersensitivity to the anti-cancer drug olaparib. Thus, our study reveals a unique mechanism mediated by KAT2A/B-dependent acetylation of a non-histone protein, which fine-tunes the DNA damage response and hence promotes genome stability.

cell biology↗

bioRxiv2018

Molecular flexibility of DNA as a major determinant of RAD51 recruitment

The timely activation of homologous recombination is essential for the maintenance of genome stability, in which the RAD51 recombinase plays a central role. Biochemically, human RAD51 polymerises faster on single-stranded DNA (ssDNA) compared to double-stranded DNA (dsDNA), raising a key conceptual question: how does it discriminate between them? In this study, we tackled this problem by systematically assessing RAD51 binding kinetics on ssDNA and dsDNA differing in length and flexibility using surface plasmon resonance. By fitting detailed polymerisation models informed by our experimental datasets, we show that RAD51 is a mechano-sensor that exhibits a larger polymerisation rate constant on flexible ssDNA compared to rigid ssDNA or dsDNA. This model presents a new general framework suggesting that the flexibility of DNA, which may increase locally as a result of DNA damage, plays an important role in rapidly recruiting repair factors that multimerise at sites of DNA damage.

biochemistry↗