Biology subjects

Mason, J. M.

Publications and source records attributed to Mason, J. M..

7 recordsLinked to original sources

bioRxiv2026

RAD54L promotes nascent DNA degradation and radial chromosome formation in FANC-deficient cells

Interstrand crosslinks are cytotoxic lesions that inhibit essential processes including replication and transcription. Replication fork reversal occurs in response to interstrand crosslink inducing drug, MMC, but how replication fork reversal promotes repair of interstrand crosslinks is poorly understood. Here, we investigated the role of the RAD54L translocase in interstrand crosslink repair. We found RAD54L is required to promote nascent DNA degradation in FANCD2 and FANCA-depleted cells consistent with a previous study indicating RAD54L promotes replication fork reversal. We further show RAD54L activity is required for formation of radial chromosomes in FANCD2-deficient cells suggesting fork reversal may be required to generate the intermediate undergoing aberrant fusion in FANC-deficient cells. Finally, we demonstrate FANCD2 foci accumulate and DSBs persist in RAD54L-deficient cells indicating RAD54L is required for efficient repair of DSBs. Together, our results indicate RAD54L plays multiple roles in efficient processing and repair of interstrand crosslinks.

molecular biology↗

bioRxiv2026

Intracellular Bismuth Coordination of Peptides and Miniproteins

Cyclisation of peptides has emerged as a powerful methodology to enhance molecular stability and functional efficacy. Bismuth(III) coordination represents a particularly compact and chemoselective cyclisation modality, yet to date has remained confined to in vitro or phage-based systems. Here we report the first example of bismuth-mediated peptide and minprotein cyclisation occurring within living cells, achieved directly during recombinant expression in E. coli. Supplementation of growth media with bismuth salts enables efficient intracellular coordination of either three or six cysteine residues, yielding bicyclic and tetracyclic architectures with minimal cellular toxicity. Bis-bismuth coordination generated a tetra-cyclic minprotein with exceptional thermal and serum stability, representing a level of intracellular structural reinforcement not previously accessible. This chemistry enables the intracellular constraint of peptides and proteins and is directly compatible with genetically encoded peptide and miniprotein libraries, establishing bismuth coordination as a compact and robust modality for live-cell molecular screening.

synthetic biology↗

bioRxiv2025

Enhancing the efficacy, utility and throughput of the Transcription Block Survival peptide library screening platform

Genetically encoded peptide library screening is a powerful strategy for discovering inhibitors of protein-protein and protein-DNA interactions. The Transcription Block Survival (TBS) assay enables the in vivo selection of peptides that antagonise transcription factor (TF) binding by linking inhibition of DNA-binding to E.coli survival. However, previous TBS implementations required laborious re-engineering of the mDHFR coding region for each new target, limiting utility. Here, we present an enhanced and streamlined TBS platform that increases throughput, simplifies target switching and improves selection stringency. By relocating TF-DNA binding sites from within the mDHFR coding sequence into the mDHFR 5-promoter/untranslated region, we preserve mDHFR folding and function, enabling rapid interchange of TF targets without the need for extensive construct redesign. We validated this system using three distinct TF targets, CREB1, ATF2 and DLX5, and two distinct consensus sites, demonstrating robust transcriptional block upon TF binding and efficient growth rescue upon peptide-mediated antagonism. Importantly, we expand the platform to accommodate full-length TFs, as exemplified by DLX5, allowing selection against biologically relevant multi-domain proteins. TBS continues to function exclusively by selecting for disruption of protein-DNA binding, ensuring mechanistic precision. Using this optimised TBS system, we successfully screened an 11.3-million-member peptide library to identify a potent antagonist of ATF2-CRE DNA binding within three months. This next generation TBS-platform dramatically improves screening efficiency and selection pressure while maintaining high biological relevance, providing a versatile and scalable tool for discovering functional peptide inhibitors of protein-DNA interactions with therapeutic potential.

synthetic biology↗

bioRxiv2025

Overexpression of the NEk8 kinase inhibits homologous recombination

Homologous recombination maintains genome stability by repairing double strand breaks and protecting replication fork stability. Defects in homologous recombination results in cancer predisposition but can be exploited due to increased sensitivity to certain chemotherapeutics such as PARP inhibitors. The NEK8 kinase has roles in the replication response and homologous recombination. NEK8 is overexpressed in breast cancer, but the impact of NEK8 overexpression on homologous recombination has not been determined. Here, we demonstrate NEK8 overexpression inhibits RAD51 focus formation resulting in a defect in homologous recombination and degradation of stalled replication forks. Importantly, NEK8 overexpression sensitizes cells to the PARP inhibitor, Olaparib. Together, our results suggest NEK8 overexpressing tumors may be recombination-deficient and respond to chemotherapeutics that target defects in recombination such as Olaparib.

cell biology↗

bioRxiv2023

UBC13-mediated template switching promotes replication stress resistance in FBH1-deficient cells

Withdrawal StatementThe authors have withdrawn this manuscript because experiments completed with more sensitive detection methods have led to results inconsistent with the original data and conclusions of the original manuscript. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.

molecular biology↗

bioRxiv2020

Single molecule imaging reveals cFos is capable of binding to and diffusing on DNA tightropes independently of cJun

AP-1 proteins are members of the basic leucine zipper (bZIP) family of dimeric transcription factors, which facilitate a multitude of cellular processes, but are primarily known for their oncogenic potential in several cancer types. The oncogenic transcription factor AP-1 binds a specific DNA target site (5TCA[G/C]TGA), however the physical mechanism of how this is achieved has not been determined. The archetypal AP-1 complex is formed by cFos and cJun, which heterodimerize via their leucine zipper domains. We investigated the DNA-binding bZIP domains of AP-1 interacting with DNA tightropes using real-time single molecule fluorescence imaging in vitro. We find that AP-1 bZIP domains comprising cFos:cJun and cJun:cJun rapidly scan DNA using a 1D diffusional search with average diffusion constants of 0.14 m2s-1 and 0.26 m2s-1 respectively. We also report for the first time that cFos is able to bind to and diffuse on DNA (0.29 m2s-1) as a mixed population of monomers and homodimers, despite previous studies suggesting that it is incapable of independent DNA binding. Additionally, we note increased pause lifetimes for the cFos:cJun heterodimer compared to the cJun:cJun homodimer, and were able to detect distinct pausing behaviours within diffusion data. Understanding how cFos:cJun and other transcription factors identify their targets is highly relevant to the development of new therapeutics which target DNA binding proteins using these search mechanisms.

biochemistry↗

bioRxiv2017

A pooled sequencing approach identifies a candidate meiotic driver in Drosophila

Meiotic drive occurs when a selfish element increases its transmission frequency above the Mendelian ratio by hijacking the asymmetric divisions of female meiosis. Meiotic drive causes genomic conflict and potentially has a major impact on genome evolution, but only a few drive loci of large effect have been described. New methods to reliably detect meiotic drive are therefore needed, particularly for discovering moderate-strength drivers that are likely to be more prevalent in natural populations than strong drivers. Here we report an efficient method that uses sequencing of large pools of backcross (BC1) progeny to test for deviations from Mendelian segregation genome-wide of single-nucleotide polymorphisms (SNPs) that distinguish the parental strains. We show that meiotic drive can be detected by a characteristic pattern of decay in distortion of SNP frequencies, caused by recombination unlinking the driver from distal loci. We further show that control crosses allow allele-frequency distortion caused by meiotic drive to be distinguished from distortion resulting from developmental effects. We used this approach to test whether chromosomes with extreme telomere-length differences segregate at Mendelian ratios, as telomeric regions are a potential hotspot for meiotic drive due to their roles in meiotic segregation and multiple observations of high rates of telomere sequence evolution. Using four different pairings of long and short telomere strains, we find no evidence that extreme telomere-length variation causes meiotic drive in Drosophila. However, we identify one candidate meiotic driver in a centromere-linked region that shows an ~8% increase in transmission frequency, corresponding to a ~54:46 segregation ratio. Our results show that candidate meiotic drivers of moderate strength can be readily detected and localized in pools of F1 progeny.

genetics↗