Biology subjects

Manjon, A. G.

Publications and source records attributed to Manjon, A. G..

6 recordsLinked to original sources

bioRxiv2025

Megabase-scale detachment of genome-nuclear lamina interactions is required for efficient repair of double stranded breaks

Lamina-associated domains (LADs) are large genomic regions that contact the nuclear lamina (NL). Double-strand breaks (DSBs) in LADs are known to be repaired more slowly and with different pathway preferences compared to other chromatin contexts. However, little is known about the chromatin changes at LADs that occur during DSB repair. Here, we report that a single DSB inside a LAD can cause detachment from the NL over several megabases. This profound spatial rearrangement is transient and reverts within 48 hours. Preventing this detachment slows down repair kinetics and renders repair incomplete, indicating that NL detachment is required for efficient repair of DSBs in LADs. NL detachment is dependent on {psi}H2AX and ATM, while it is antagonized by DNAPKcs activity. Remarkably, {psi}H2AX also antagonizes NL interactions at chromosome ends. Taken together, our data indicate that {psi}H2AX accumulation in LADs induces large scale rewiring of genome-NL interactions, allowing for efficient repair of DSBs.

cell biology↗

bioRxiv2022

Widespread chromatin context-dependencies ofDNA double-strand break repair proteins

DNA double-strand breaks are repaired by multiple pathways, including non-homologous end-joining (NHEJ) and microhomology-mediated end-joining (MMEJ). The balance of these pathways is dependent on the local chromatin context, but the underlying mechanisms are poorly understood. By combining knockout screening with a dual MMEJ:NHEJ reporter inserted in 19 different chromatin environments, we identified dozens of DNA repair proteins that modulate pathway balance dependent on the local chromatin state. Proteins that favor NHEJ mostly synergize with euchromatin, while proteins that favor MMEJ generally synergize with distinct types of heterochromatin. BRCA2 is an example of the former, which is corroborated by chromatin-dependent shifts in mutation patterns of BRCA2-/- cancer genomes. These results uncover a complex network of proteins that regulate MMEJ:NHEJ balance in a chromatin context-dependent manner. ONE SENTENCE SUMMARYA multiplexed screen reveals how dozens of proteins sense the local chromatin context to tune the balance between two DNA repair pathways.

molecular biology↗

bioRxiv2021

Unexpected gene activation following CRISPR-Cas9-mediated genome editing

The discovery of the Clustered Regularly-Interspaced Short Palindromic Repeats (CRISPR) and its development as a genome editing tool has revolutionized the field of molecular biology. In the DNA damage field, CRISPR has brought an alternative to induce endogenous double-strand breaks (DSB) at desired genomic locations and study the DNA damage response and its consequences. Many systems for sgRNA delivery have been reported in order to efficiency generate this DSB, including lentiviral vectors. However, some of the consequences of these systems are yet not well understood. Here we report that lentiviral-based sgRNA vectors can integrate into the endogenous genomic target location, leading to undesired activation of the target gene. By generating a DSB in the regulatory region of the ABCB1 gene using a lentiviral sgRNA vector, we can induce the formation of taxol-resistant colonies. We show that these colonies upregulated ABCB1 via integration of the EEF1A1 and the U6 promoters from the sgRNA vector. We believe that this is an unreported CRISPR/Cas9 artefact that researchers need to be aware of when using lentiviral vectors for genome editing.

genomics↗

bioRxiv2021

Perturbations in 3D genome organization can promote acquired drug resistance

Acquired drug resistance is a major problem in the treatment of cancer. hTERT-immortalized, untransformed RPE-1 (RPE) cells can acquire resistance to taxol by derepressing the ABCB1 gene, encoding for the multidrug transporter P-gP. Here we have investigated how the ABCB1 gene is derepressed. We show that activation of the ABCB1 gene is associated with reduced DNA methylation, reduced H3K9 trimethylation and increased H3K27 acetylation at the ABCB1 promoter. In addition, we find that the ABCB1 locus has moved away from the nuclear lamina in the taxol-resistant cells. This raises the question which of these alterations were causal to derepression. Directly modifying DNA methylation or H3K27 methylation had neither significant effect on ABCB1 expression, nor did it promote drug resistance. In contrast, the disruption of Lamin B Receptor (LBR), a component of the nuclear lamina involved in genome organization, did promote the acquisition of a taxol-resistant phenotype in a subset of cells. Using CRISPRa-mediated gene activation, we could further substantiate a model in which disruption of lamina association renders the ABCB1 gene permissive to derepression. Based on these data we propose a model in which nuclear lamina dissociation of a repressed gene allows for its activation, implying that deregulation of the 3D genome topology could play an important role in tumor evolution and the acquisition of drug resistance.

molecular biology↗

bioRxiv2020

PRECISE+ predicts drug response in patients by non-linear subspace-based transfer from cell lines and PDX models

Pre-clinical models have been the workhorse of cancer research for decades. While powerful, these models do not fully recapitulate the complexity of human tumors. Consequently, translating biomarkers of drug response from pre-clinical models to human tumors has been particularly challenging. To explicitly take these differences into account and enable an efficient exploitation of the vast pre-clinical drug response resources, we developed TRANSACT, a novel computational framework for clinical drug response prediction. First, TRANSACT employs non-linear manifold learning to capture biological processes active in pre-clinical models and human tumors. Then, TRANSACT builds predictors on cell line response only and transfers these to Patient-Derived Xenografts (PDXs) and human tumors. TRANSACT outperforms four competing approaches, including Deep Learning approaches, for a set of 15 drugs on PDXs, TCGA cohorts and 226 metastatic tumors from the Hartwig Medical Foundation data. For only four drugs Deep Learning outperforms TRANSACT. We further derived an algorithmic approach to interpret TRANSACT and used it to validate the approach by identifying known biomarkers to targeted therapies and we propose novel putative biomarkers of resistance to Paclitaxel and Gemcitabine.

bioinformatics↗

bioRxiv2019

DNA end-resection in highly accessible chromatin produces a toxic break

The authors have withdrawn their manuscript whilst they perform additional experiments to test some of their conclusions further. Despite repetitive attempts to reproduce the data shown in figure 2I, we have failed to obtain convincing evidence that chemical inhibition of Mre11 can improve the fitness of cells exposed to the so-called "halt" breaks. Upon re-analysis of the raw data used to compose figure 2I, we noted inconsistencies in the inclusion of datapoints, for which a satisfactory motivation was not apparent. In order to reproduce the experiments with the DNA-PK inhibitor, we ordered new batches of this compound, but noticed that the IC50 was significantly lower than that of the earlier batch (as determined in suppl. fig.1). While this might be due to a difference in batches, the IC50 of the newer batches was more comparable to the IC50 of this compound described for other cell lines. This sheds doubts on the quality of the batch used for the experiments shown in our manuscript, and we no longer wish to draw any conclusions based on these experiments. The authors hope that future experiments can resolve if open chromatin does indeed produce a potentially more toxic DNA break. 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.

cell biology↗