Factors That Affect the Formation of Chromosomal Translocations in Cells

Chromosomal translocations are products of the illegitimate repair of DNA double-strand breaks (DSBs). Their formation can bring about significant structural and molecular changes in the cell that can be physiologically and pathologically relevant. The induced changes may lead to serious and life-threatening diseases such as cancer. As a growing body of evidence suggests, the formation of chromosomal translocation is not only affected by the mere close spatial proximity of gene loci as potential translocation partners. Several factors may affect formation of chromosomal translocations, including chromatin motion to the potential sources of DSBs in the cell. While these can be apparently random events, certain chromosomal translocations appear to be cell-type-specific. In this review, we discuss how chromosomal translocations are formed and explore how different cellular factors contribute to their formation

Interaction between mesenchymal stem cells and myoblasts in the context of facioscapulohumeral muscular dystrophy contributes to the disease phenotype

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Exchange of subtelomeric regions between chromosomes 4q and 10q reverts the FSHD genotype and phenotype

International audienceThe most common form of facioscapulohumeral dystrophy (FSHD1) is caused by a partial loss of the D4Z4 macrosatellite repeat array in the subtelomeric region of chromosome 4. Patients with FSHD1 typically carry 1 to 10 D4Z4 repeats, whereas nonaffected individuals have 11 to 150 repeats. The ~150-kilobyte subtelomeric region of the chromosome 10q exhibits a ~99% sequence identity to the 4q, including the D4Z4 array. Nevertheless, contractions of the chr10 array do not cause FSHD or any known disease, as in most people D4Z4 array on chr10 is flanked by the nonfunctional polyadenylation signal, not permitting the DUX4 expression. Here, we attempted to correct the FSHD genotype by a CRISPR-Cas9–induced exchange of the chr4 and chr10 subtelomeric regions. We demonstrated that the induced t(4;10) translocation can generate recombinant genotypes translated into improved FSHD phenotype. FSHD myoblasts with the t(4;10) exhibited reduced expression of the DUX4 targets, restored PAX7 target expression, reduced sensitivity to oxidative stress, and improved differentiation capacity

Urokinase Receptor uPAR Downregulation in Neuroblastoma Leads to Dormancy, Chemoresistance and Metastasis

uPAR is a membrane receptor that binds extracellular protease urokinase, contributes to matrix remodeling and plays a crucial role in cellular adhesion, proliferation, survival, and migration. uPAR overexpression in tumor cells promotes mitogenesis, opening a prospective avenue for targeted therapy. However, uPAR targeting in cancer has potential risks. We have recently shown that uPAR downregulation in neuroblastoma promotes epithelial-mesenchymal transition (EMT), potentially associated with metastasis and chemoresistance. We used data mining to evaluate the role of uPAR expression in primary and relapsed human neuroblastomas. To model the decreased uPAR expression, we targeted uPAR using CRISPR/Cas9 and shRNA in neuroblastoma Neuro2a cells and evaluated their chemosensitivity in vitro as well as tumor growth and metastasis in vivo. We demonstrate that the initially high PLAUR expression predicts poor survival in human neuroblastoma. However, relapsed neuroblastomas have a significantly decreased PLAUR expression. uPAR targeting in neuroblastoma Neuro2a cells leads to p38 activation and an increased p21 expression (suggesting a dormant phenotype). The dormancy in neuroblastoma cells can be triggered by the disruption of uPAR-integrin interaction. uPAR-deficient cells are less sensitive to cisplatin and doxorubicin treatment and exhibit lower p53 activation. Finally, low uPAR-expressing Neuro2a cells formed smaller primary tumors, but more frequent metastasis in mice. To the best of our knowledge, this is the first study revealing the pathological role of dormant uPAR-deficient cancer cells having a chemoresistant and motile phenotype

Novel Fluorescent Mitochondria-Targeted Probe MitoCLox Reports Lipid Peroxidation in Response to Oxidative Stress In Vivo

A new mitochondria-targeted probe MitoCLox was designed as a starting compound for a series of probes sensitive to cardiolipin (CL) peroxidation. Fluorescence microscopy reported selective accumulation of MitoCLox in mitochondria of diverse living cell cultures and its oxidation under stress conditions, particularly those known to cause a selective cardiolipin oxidation. Ratiometric fluorescence measurements using flow cytometry showed a remarkable dependence of the MitoCLox dynamic range on the oxidation of the sample. Specifically, MitoCLox oxidation was induced by low doses of hydrogen peroxide or organic hydroperoxide. The mitochondria-targeted antioxidant 10-(6-plastoquinonyl)decyltriphenyl-phosphonium (SkQ1), which was shown earlier to selectively protect cardiolipin from oxidation, prevented hydrogen peroxide-induced MitoCLox oxidation in the cells. Concurrent tracing of MitoCLox oxidation and membrane potential changes in response to hydrogen peroxide addition showed that the oxidation of MitoCLox started without a delay and was complete during the first hour, whereas the membrane potential started to decay after 40 minutes of incubation. Hence, MitoCLox could be used for splitting the cell response to oxidative stress into separate steps. Application of MitoCLox revealed heterogeneity of the mitochondrial population; in living endothelial cells, a fraction of small, rounded mitochondria with an increased level of lipid peroxidation were detected near the nucleus. In addition, the MitoCLox staining revealed a specific fraction of cells with an increased level of oxidized lipids also in the culture of human myoblasts. The fraction of such cells increased in high-density cultures. These specific conditions correspond to the initiation of spontaneous myogenesis in vitro, which indicates that oxidation may precede the onset of myogenic differentiation. These data point to a possible participation of oxidized CL in cell signalling and differentiation

Urokinase System in Pathogenesis of Pulmonary Fibrosis: A Hidden Threat of COVID-19

Pulmonary fibrosis is a common and threatening post-COVID-19 complication with poorly resolved molecular mechanisms and no established treatment. The plasminogen activator system, including urokinase (uPA) and urokinase receptor (uPAR), is involved in the pathogenesis of COVID-19 and contributes to the development of lung injury and post-COVID-19 pulmonary fibrosis, although their cellular and molecular underpinnings still remain obscure. The aim of the current study was to assess the role of uPA and uPAR in the pathogenesis of pulmonary fibrosis. We analyzed uPA and uPAR expression in human lung tissues from COVID-19 patients with pulmonary fibrosis using single-cell RNA-seq and immunohistochemistry. We modeled lung fibrosis in Plau-/- and Plaur-/- mice upon bleomycin instillation and explored the effect of uPAR downregulation in A549 and BEAS-2B lung epithelial cells. We found that uPAR expression drastically decreased in the epithelial airway basal cells and monocyte/macrophage cells, whereas uPA accumulation significantly increased in tissue samples of COVID-19 patients. Lung injury and fibrosis in Plaur-/- vs. WT mice upon bleomycin instillation revealed that uPAR deficiency resulted in pro-fibrogenic uPA accumulation, IL-6 and ACE2 upregulation in lung tissues and was associated with severe fibrosis, weight loss and poor survival. uPAR downregulation in A549 and BEAS-2B was linked to an increased N-cadherin expression, indicating the onset of epithelial–mesenchymal transition and potentially contributing to pulmonary fibrosis. Here for the first time, we demonstrate that plasminogen treatment reversed lung fibrosis in Plaur-/- mice: the intravenous injection of 1 mg of plasminogen on the 21st day of bleomycin-induced fibrosis resulted in a more than a two-fold decrease in the area of lung fibrosis as compared to non-treated mice as evaluated by the 42nd day. The expression and function of the plasminogen activator system are dysregulated upon COVID-19 infection, leading to excessive pulmonary fibrosis and worsening the prognosis. The potential of plasminogen as a life-saving treatment for non-resolving post-COVID-19 pulmonary fibrosis warrants further investigation

T‐cell‐derived Hodgkin lymphoma has motility characteristics intermediate between Hodgkin and anaplastic large cell lymphoma

International audienceAbstract Imbalance in the finely orchestrated system of chromatin-modifying enzymes is a hallmark of many pathologies such as cancers, since causing the affection of the epigenome and transcriptional reprogramming. Here, we demonstrate that a loss-of-function mutation (LOF) of the major histone lysine methyltransferase SETDB1 possessing oncogenic activity in lung cancer cells leads to broad changes in the overall architecture and mechanical properties of the nucleus through genome-wide redistribution of heterochromatin, which perturbs chromatin spatial compartmentalization. Together with the enforced activation of the epithelial expression program, cytoskeleton remodeling, reduced proliferation rate and restricted cellular migration, this leads to the reversed oncogenic potential of lung adenocarcinoma cells. These results emphasize an essential role of chromatin architecture in the determination of oncogenic programs and illustrate a relationship between gene expression, epigenome, 3D genome and nuclear mechanics

SETDB1 fuels the lung cancer phenotype by modulating epigenome, 3D genome organization and chromatin mechanical properties

International audienceAbstract Imbalance in the finely orchestrated system of chromatin-modifying enzymes is a hallmark of many pathologies such as cancers, since causing the affection of the epigenome and transcriptional reprogramming. Here, we demonstrate that a loss-of-function mutation (LOF) of the major histone lysine methyltransferase SETDB1 possessing oncogenic activity in lung cancer cells leads to broad changes in the overall architecture and mechanical properties of the nucleus through genome-wide redistribution of heterochromatin, which perturbs chromatin spatial compartmentalization. Together with the enforced activation of the epithelial expression program, cytoskeleton remodeling, reduced proliferation rate and restricted cellular migration, this leads to the reversed oncogenic potential of lung adenocarcinoma cells. These results emphasize an essential role of chromatin architecture in the determination of oncogenic programs and illustrate a relationship between gene expression, epigenome, 3D genome and nuclear mechanics