Investigating the role of RNA-binding proteins in telomere replication stress and alternative lengthening of telomeres

Telomeres are protective structures at chromosome ends that prevent genome instability. In most cancer cells, telomere shortening is counteracted by telomerase reactivation. However, 10–15% of cancers, including aggressive types like osteosarcoma and glioblastoma, use a telomerase-independent pathway known as Alternative Lengthening of Telomeres (ALT). ALT relies on homology-directed DNA repair and is triggered by persistent replication stress at telomeres. Despite recent advancements in cancer therapy, ALT+ cancers currently lack targeted therapies and have a poor prognosis, highlighting the need to increase our understanding of ALT and develop new treatments against these aggressive tumours.Increasing evidence suggests a role of RNA-binding proteins (RBPs) in DNA replication stress response and telomere stability. Therefore, I hypothesise that a subset of RBPs influence telomere replication stress and can modulate ALT activity. Towards this, I plan to:1) Perform a CRISPR-based screen in ALT-positive and ALT-negative cancer cell lines to identify RBPs that affect telomeric replication stress and ALT activity.2) Study how top candidate RBPs regulate telomere replication stress and ALT-specific telomere DNA synthesis using a range of molecular assays.3) Evaluate the therapeutic potential of targeting these RBPs in ALT-positive cancers by testing their effects on cell viability and response to DNA damaging agents.This work will reveal new RBP regulators of ALT and provide insight into the interplay between RNA metabolism, replication stress, and telomere maintenance. This project will provide a deeper, mechanistic understanding of how RNA-binding proteins influence telomere replication stress and ALT, filling a critical gap in telomere biology and paving the way for innovative cancer treatments against these aggressive tumours.