RADX Sustains POT1 Function at Telomeres to Counteract RAD51 Binding, which Triggers Telomere Fragility

Summary The 3’ terminal DNA extensions at chromosome ends can become engaged into multiple biochemical reactions during DNA replication, telomerase-mediated telomere extension, homology directed DNA repair, nucleolytic processing and DNA damage checkpoint activation. To keep these activities in check, telomeric 3’ overhangs can be hidden in t-loop structures or they associate with specialized proteins such as POT1. Here, we explore the telomeric microenvironment using a proximity-dependent labeling approach and identify the oligonucleotide/oligosaccharide-binding (OB)-fold containing protein RADX. RADX binds single-stranded telomeric DNA throughout the cell cycle along with POT1, suppressing accumulation of fragile telomeres, which are indicative of telomere replication defects. Telomere fragility in POT1 and RADX double-depleted cells was due to accumulation of the RAD51 recombinase at telomeres. RADX also supports DNA damage signaling at POT1-depleted telomeres counteracting RAD51 binding. Thus, RADX represents next to POT1 a second OB-fold containing single-strand telomere binding protein sustaining telomere protection.