Biology subjects

Ragu, S.

Publications and source records attributed to Ragu, S..

3 recordsLinked to original sources

bioRxiv2025

Dysregulation of replication stress-induced ROS in transformed cell lines: a vicious circle at cancer initiation

The canonical DNA damage response (DDR) maintains genome stability, involving DNA synthesis/cell cycle arrest. However, unchallenged cells proliferate when they are continually exposed to low-level/endogenous replication stress. We previously discovered and characterized a noncanonical cellular response that is specific to nonblocking replication stress, i.e., low-level stress (LoL-DDR), in primary cells. Although this response generates replication stress-induced reactive oxygen species (RIR), it triggers a program that prevents the accumulation of premutagenic 8-oxo-guanine (8-oxoG). Primary cells control RIR production via NADPH oxidases. Increasing the severity of replication stress above a precise threshold triggers the canonical DDR, leading to cell cycle arrest and RIR suppression, resulting in a peak-shaped dose response for RIR production. Here, we show that the LoL-DDR is dysregulated in cancer cell lines, which exhibit the following differences compared with primary cells: 1-RIR are not detoxified under high-level stress conditions, resulting in a continuous increase in the dose-response curve of RIR production; 2-RIR are not produced by NADPH oxidases; and 3-replication stress favors the accumulation of the premutagenic 8-oxoG. Moreover, using an in vitro breast cancer progression model, we show that LoL-DDR dysregulation occurs at an early stage of cancer progression. Since, conversely, ROS trigger replication stress this establishes a "vicious circle" involving replication-stress and ROS that continuously jeopardizes genome integrity that should fuel and amplify tumorigenesis.

cell biology↗

bioRxiv2020

An adaptive pre-DNA-damage-response protects genome integrity

The DNA damage response (DDR) interrupts cell cycle progression to restore genome integrity. However, unchallenged proliferating cells are continually exposed to endogenous stress, raising the question of a stress-threshold for DDR activation. Here, we identified a stress threshold below which primary human fibroblasts, activate a cell-autonomous response that not activates full DDR and not arrests cell cycle progression. We characterized this "pre-DDR" response showing that it triggers the production of reactive oxygen species (ROS) by the NADPH oxidases DUOX1 and DUOX2, under the control of NF-{kappa}B and PARP1. Then, replication stress-induced ROS (RIR) activates the FOXO1 detoxifying pathway, preventing the nuclear accumulation of the pre-mutagenic 8-oxoGuanine lesion, upon endogenous as well as exogenous pro-oxidant stress. Increasing the replication stress severity above the threshold triggers the canonical DDR, leading to cell cycle progression arrest, but also to RIR suppression. These data reveal that cells adapt their response to stress severity, unveiling a tightly regulated "pre-DDR" adaptive response that protects genome integrity without arresting cell cycle progression.

cell biology↗

bioRxiv2019

Mammalian RAD51 prevents non-conservative alternative end-joining and single strand annealing through non-catalytic mechanismssms

The selection of the DNA double-strand breaks (DSBs) repair pathway is decisive for genetic stability/instability. We proposed that it acts according to two successive steps: 1-canonical non-homologous end-joining (C-NHEJ) versus single-strand DNA (ssDNA) resection; 2- on ssDNA, gene conversion (GC) versus non-conservative single-strand annealing (SSA) or alternative end-joining (A-EJ).\n\nUsing intramolecular substrates, we systematically analysed the equilibrium between the different DSB repair pathways. We show that ablation of RAD51 stimulated both SSA and A-EJ but did not stimulate C-NHEJ, validating the two-step model. Moreover, we found that two ATP-mutant dominant-negative forms of RAD51 that stimulated non-conservative repair, failed to load into damaged chromatin, clarifying the role of ATP in RAD51-mediated HR, also. In contrast, another dominant-negative form of RAD51, which retains its DNA binding capacities, repressed SSA and A-EJ, revealing two separable functions of RAD51 i.e. GC and non-conservative repair inhibition. In vitro assays show that the binding of RAD51 on both complementary ssDNA is required to block both spontaneous and RAD52-induced strand annealing. Therefore, RAD51 represses non-conservative repair (SSA and A-EJ), by inhibiting the annealing step through ssDNA occupancy, independently of the catalytic strand-exchange activity required for GC.

molecular biology↗