The BLM DHBN domain and structure-selective nucleases drive mitotic arrest-dependent telomere deprotection under WRN and TRF2 control

Telomeres shield chromosome ends from DNA damage response through T-loops, lariat DNA structures formed and stabilized by the shelterin protein TRF2. During prolonged mitotic arrest, telomeres lose this protection through a process termed mitotic arrest-dependent (MAD) telomere deprotection, which elicits telomere-specific DNA damage signaling in the absence of telomere shortening or chromosome end-to-end fusions. We previously demonstrated that the RecQ helicase BLM promotes MAD telomere deprotection, whereas the related helicase WRN suppresses it independently of its catalytic activities. However, the molecular interplay between BLM and WRN at mitotic telomeres and whether additional recombination-associated enzymes contribute to MAD telomere deprotection have remained unresolved. Here, we identify the Dimerization Helical Bundle in the N-terminal (DHBN) domain of BLM helicase as the critical determinant of MAD telomere deprotection and show that WRN selectively restrains this activity without interfering with BLM's canonical genome-protective functions. We further show that both MUS81 and GEN1 contribute to MAD telomere deprotection. Moreover, the exacerbation of MAD telomere deprotection observed upon TRF2 depletion is strongly attenuated by additional depletion of these enzymes, demonstrating that TRF2 normally protects T-loop junctions from their enzymatic activities. Collectively, our findings reveal how telomeres become selectively vulnerable during prolonged mitotic arrest and uncover a regulated enzymatic mechanism that repurposes recombination machinery at chromosome ends.