Abstract 4198: Unraveling the molecular interactions between DDR and autophagy in response to oxidative stress

Abstract The DNA damage response (DDR) is essential for maintaining genomic integrity by detecting and repairing DNA damage. In conjunction with autophagy, DDR functions as a critical protective mechanism under oxidative stress, mitigating the risk of cancer development. Our study investigates the link between oxidative stress and DDR, particularly focusing on ATM kinase's role in reactive oxygen species (ROS)-mediated genotoxic stress for double-strand break (DSB) repair. Cancer cells' heightened reliance on DDR, due to high proliferation and genomic instability, underscores their susceptibility to DDR inhibitors. However, the effectiveness of DDR inhibitors is often limited by the evolution of resistance in tumors. Resistance mechanisms can include the restoration of DNA repair pathways or activation of compensatory pathways. This adaptability limits the long-term efficacy of DDR-targeted therapies, emphasizing the need to understand the molecular drivers underlying resistance to enable the development of more effective strategies. Our study focuses on the potential interaction between Mediator of DNA damage checkpoint 1 (MDC1), a key DDR protein, and Beclin-1, a vital autophagy inducer. We have employed a comprehensive approach using the HeLa cell line, including wild type, MDC1 knockdown (KD), and MDC1 overexpressing (MDC1-Ov) variants. Oxidative stress was induced using hydrogen peroxide (H2O2), and we conducted co-immunoprecipitation, immunofluorescence, and immunoblotting to investigate the interactions between MDC1 and Beclin-1 under these conditions. Our results demonstrate that MDC1 interacts with Beclin-1, with the interaction enhanced over time, indicating its essential role in cellular responses to oxidative stress. We also show that oxidative stress prompts the relocation of Beclin-1 from the cytoplasm to the nucleus, suggesting that Beclin-1 may be involved in regulating DDR in response to oxidative stress-induced DNA damage. Given the central role of MDC1 in recruiting various effector proteins at the site of DNA repair, we hypothesized that MDC1 may facilitate the nuclear translocation of Beclin-1. Consistent with this hypothesis, we found that nuclear localization of Beclin-1 was significantly reduced in MDC1 knockdown cells compared to wild type when treated with H2O2. These findings underscore the critical role of MDC1 in modulating the nuclear localization of Beclin-1 in response to oxidative stress. The significance of our study lies in its potential implications for chemotherapy. Given the prevalence of oxidative stress in various cancers and the therapeutic reliance on chemotherapy-induced oxidative stress, understanding this novel interplay between DDR and autophagy pathways may provide insights for developing constructive therapeutic strategies. Citation Format: Kavya Ajit Pandya, Neeru Singh. Unraveling the molecular interactions between DDR and autophagy in response to oxidative stress [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4198.