Biology subjects

Benedetto, S.

Publications and source records attributed to Benedetto, S..

2 recordsLinked to original sources

bioRxiv2025

Recurrent DNA Break Clusters Regulated by Polymerase Theta Are Essential for Replication Stress Induced Copy Number Variation

Copy number variations (CNVs) are a form of genetic alteration strongly implicated in numerous neurological and psychiatric disorders, as well as brain cancer. Replication stress is a common cause of CNVs. Despite the prevailing model that CNVs arise from DNA double strand breaks (DSBs), there has been no assay that directly perturbs presumed DSB sources and measures CNV output. Here, we identified a subset of recurrent DNA break clusters (RDCs) as a causal factor for CNV formation. In murine neural progenitor cells under replication stress, mapping the formation of CNVs revealed their location in RDC regions that contain actively transcribed genes. CRISPR/Cas9-mediated transcriptional suppression abrogated both RDC and CNV formation, but does not alter their replication timing. We found that DNA polymerase theta (Pol {theta}), a protector against CNV formation, plays a critical but context dependent role upstream of RDC formation. Chemically inhibiting the activity of Pol {theta} reduced end filling and micro-homology-mediated end joining in XRCC4/P53-deficient cells. Conversely, Pol {theta} inhibition led to elevated DSB density detection at RDC-containing loci in wild-type neural stem and progenitor cells, suggesting its role in preventing transcription-replication conflicts. Our data identify RDCs as contributors to genomic heterogeneity with plausible downstream effects on brain disorders and malignancy.

molecular biology↗

bioRxiv2025

Adaptive Clonal Expansion Shapes Brain Development

Embryonic neural stem/progenitor cells (NSPCs) exhibit remarkable proliferative plasticity, allowing them to fully recover neuronal populations even after substantial cell loss 1,2. However, it remains unclear whether all embryonic NSPCs respond to brain lesions. To address this, we developed a mouse model to investigate NSPC proliferation dynamics, hypothesizing that the loss of progenitor cells would induce fitness competition among NSPCs. In this model, half of the founder NSPCs were ablated using diphtheria toxin A at the onset of neurogenesis, yet the surviving cells regenerated a brain containing all neuronal types within five of the total twenty embryonic days. Analysis of allelic variants revealed overrepresented somatic variants, indicating that only a small fraction of NSPCs underwent significant clonal expansion during early neurogenesis. Modelling proliferation dynamics predicted that as few as 10% of NSPCs could produce 83% of neurons by the time of birth. Single nucleotide substitution analysis suggested a potential link to oxidative metabolism in some of the expanded clones. Moreover, single-cell transcriptomics showed delayed development and a reduced NSPC pool as consequences of adaptive clonal expansion. Our findings highlight that NSPC exhibit varying expansion potential and that adaptive clonal expansion indirectly altered neuronal cell composition in the brain.

developmental biology↗