Biology subjects

Reuveni, E.

Publications and source records attributed to Reuveni, E..

6 recordsLinked to original sources

bioRxiv2026

Mapping chromothripsis across human cancer cell lines reveals genome evolution and therapeutic vulnerabilities

Chromothripsis is a catastrophic chromosomal rearrangement found in ~40% of primary tumors, yet its landscape in cancer cell lines (CCLs) remains uncharacterized. To address this gap, we developed a pipeline to detect chromothripsis-like events from copy-number data alone, and mapped their occurrence across 1,646 human CCLs. Chromothripsis patterns were broadly conserved between primary tumors and CCLs, but CCLs displayed some distinct features and a significantly higher prevalence of chromothripsis. Chromothripsis continued to emerge during culture propagation, likely contributing to its increased prevalence in CCLs. Integrating genomic and functional data revealed selective vulnerabilities: chromothriptic cells were resistant to DNA damage-inducing agents but highly sensitive to histone deacetylase inhibitors (HDACi). Using isogenic models, we validated that HDACi could synergistically sensitize chromothriptic cells to DNA damage-inducing agents. Our study establishes the first comprehensive resource of chromothripsis in CCLs and reveals functional consequences and therapeutic vulnerabilities of chromothripsis-positive cancers.

cancer biology↗

bioRxiv2025

Expanding single-cell toolbox with a cost-effective full-length total RNA droplet-based sequencing technology

sc-rDSeq is a scalable, full-length total RNA droplet-based technology that captures both polyadenylated and non-polyadenylated RNAs, including histone-RNAs, small and long non-coding RNAs, and enhancer RNAs. It achieves a tenfold increase in UMIs per cell over inDrops while remaining simple and cost-efficient. Applied to lung cancer cells, sc-rDSeq revealed hidden heterogeneity, divergent signaling pathways, and non-polyA RNA variations undetectable with 3 end-based methods. In EGFR inhibitor-treated persister cells, it confirmed drug-induced cell cycle arrest based on histone mRNA expression, identified distinct resistance strategies. It also detected alternative splicing events and enabled single cell genotyping of SNVs linked to drug resistance. While SNVs act as drivers of drug resistance, additional layers of expression-based cellular states must be explored to fully understand and combat persister cells. These findings highlight the potential of sc-rDSeq to advance transcriptomic studies and personalized medicine by providing a comprehensive and scalable approach to cellular RNA analysis.

genetics↗

bioRxiv2023

Human aneuploid cells depend on the RAF/MEK/ERK pathway for overcoming increased DNA damage

Aneuploidy is a hallmark of human cancer, yet the cellular mechanisms that allow cells to cope with aneuploidy-induced cellular stresses remain largely unknown. Such coping mechanisms may present cellular vulnerabilities that can be harnessed for targeting cancer cells. Here, we induced aneuploidy in non-transformed RPE1-hTERT cells and derived multiple stable clones with various degrees of chromosome imbalances. We performed an unbiased genomic profiling of 6 isogenic clones, using whole-exome and RNA sequencing. We then functionally interrogated their cellular dependency landscapes, using genome-wide CRISPR/Cas9 screens and large-scale drug screens. We found that aneuploid clones activated the DNA damage response (DDR), and were consequently more resistant to further DNA damage induction. Interestingly, aneuploid cells also exhibited elevated RAF/MEK/ERK pathway activity, and were more sensitive to several clinically-relevant drugs targeting this pathway, and in particular to genetic and chemical CRAF inhibition. CRAF activity was functionally linked to the resistance to DNA damage induction, as CRAF inhibition sensitized aneuploid cells to DNA damage-inducing chemotherapies. The association between aneuploidy, RAF/MEK/ERK signaling, and DDR was independent of p53. The increased activity and dependency of aneuploid cells on the RAF/MEK/ERK pathway was validated in another isogenic aneuploid system, and across hundreds of human cancer cell lines, confirming their relevance to human cancer. Overall, our study provides a comprehensive resource for genetically-matched karyotypically-stable cells of various aneuploidy states, and reveals a novel therapeutically-relevant cellular dependency of aneuploid cells.

cancer biology↗

bioRxiv2023

Increased RNA and protein degradation is required for counteracting transcriptional burden and proteotoxic stress in human aneuploid cells

Aneuploidy, an abnormal chromosome composition, results in a stoichiometric imbalance of protein complexes, which jeopardizes the fitness of aneuploid cells. Aneuploid cells thus need to compensate for the imbalanced DNA levels by regulating their RNA and protein levels, a phenomenon known as dosage compensation. However, the molecular mechanisms involved in dosage compensation in human cells - and whether they can be targeted to selectively kill aneuploid cancer cells - remain unknown. Here, we addressed this question via molecular dissection of multiple diploid vs. aneuploid cell models. Using genomic and functional profiling of a novel isogenic system of RPE1-hTERT cells with various degrees of aneuploidy, we found that aneuploid cells cope with both transcriptional burden and proteotoxic stress. At the mRNA level, aneuploid cells increased RNA synthesis, but concomitantly elevated several RNA degradation pathways, in particular the nonsense-mediated decay (NMD) and the microRNA-mediated mRNA silencing pathways. Consequently, aneuploid cells were more sensitive to the genetic or chemical perturbation of several key components of these RNA degradation pathways. At the protein level, aneuploid cells experienced proteotoxic stress, resulting in reduced translation and increased protein degradation, rendering them more sensitive to proteasome inhibition. These findings were recapitulated across hundreds of human cancer cell lines and primary tumors, confirming that both non-transformed and transformed cells alter their RNA and protein metabolism in order to adapt to the aneuploid state. Our results reveal that aneuploid cells are dependent on the over- or under-activation of several nodes along the gene expression process, identifying these pathways as clinically-actionable vulnerabilities of aneuploid cells.

cancer biology↗

bioRxiv2022

Acute expression of human APOBEC3B in mice causes lethality associated with RNA editing

RNA editing has been described to promote heterogeneity leading to the development of multiple disorders including cancer. The cytosine deaminase APOBEC3B is known to fuel tumor evolution through DNA mutagenesis, but whether it may also function as an RNA editing enzyme has not been studied. Here, we engineered a novel doxycycline-inducible mouse model of human APOBEC3B-overexpression to understand the impact of this enzyme in tissue homeostasis and address a potential role in C-to-U RNA editing. Elevated and sustained levels of APOBEC3B led to rapid alteration of cellular fitness, major organ dysfunction, and ultimately lethality in mice. Importantly, extensive analyses of RNA-sequencing and WES from mouse tissues expressing high APOBEC3B levels reveal frequent UCC-to-UUC RNA editing events mainly localized in a specific hotspot. This work identifies, for the first time, a new function for APOBEC3B in RNA editing and presents a valuable preclinical tool to understand the emerging role of APOBEC3B as a potent driver of cancer and other diseases.

molecular biology↗

bioRxiv2021

Frequent Aneuploidy in Primary Human T Cells Following CRISPR-Cas9 cleavage

Multiple ongoing clinical trials use site-specific nucleases to disrupt T cell receptor (TCR) genes in order to allow for allogeneic T cell therapy1-5. In particular, the first U.S. clinical trial using CRISPR-Cas9 entailed the targeted disruption of the TCR chains and programmed cell death protein 1 (PDCD1) in T cells of refractory cancer patients6. Here, we used the same guide RNA sequences and applied single-cell RNA sequencing (scRNAseq) to more than 7000 primary human T cells, transfected with CRISPR-Cas9. Four days post-transfection, we found a loss of chromosome 14, harboring the TCR locus, in up to 9% of the cells, and a chromosome 14 gain in up to 1.4% of the cells. We further identified truncations of chromosome 7, harboring the TCR{beta} locus, in 9.9% of the cells. Loss of heterozygosity (LOH) was further validated using fluorescence in situ hybridization (FISH) and the temporal dynamics of cleavage and incomplete repair were monitored using digital droplet PCR (ddPCR). Aneuploidy was found among all T cell subsets and was associated with transcriptional signatures of reduced proliferation and metabolism as well as with induced p53 activation and cell death. We conclude that aneuploidy and chromosomal truncations are frequent outcomes of CRISPR-Cas9 cleavage in clinical protocols. Monitoring and minimizing these aberrant products is crucial for future applications of genome editing in T cell engineering and beyond.

bioengineering↗