The DNA Replication Checkpoint Targets the Kinetochore for Relocation of Collapsed Forks to the Nuclear Periphery

Hairpin forming expanded CAG/CTG repeats pose significant challenges to DNA replication which can lead to replication fork collapse. Long CAG/CTG repeat tracts relocate to the nuclear pore complex to maintain their integrity. Forks impeded by DNA structures are known to activate the DNA damage checkpoint, thus we asked whether checkpoint proteins play a role in relocation of collapsed forks to the nuclear periphery in S. cerevisiae . We show that relocation of a (CAG/CTG) ₁₃₀ tract is dependent on activation of the Mrc1/Rad53 replication checkpoint. Further, checkpoint-mediated phosphorylation of the kinetochore protein Cep3 is required for relocation, implicating detachment of the centromere from the spindle pole body. Activation of this pathway leads to DNA damage-induced microtubule recruitment to the repeat. These data suggest a role for the DNA replication checkpoint in facilitating movement of collapsed replication forks to the nuclear periphery by centromere release and microtubule-directed motion.

The DNA replication checkpoint initiates relocation of a structure-forming CAG repeat tract to the nuclear pore complex (NPC)The importance of Mrc1 (hClaspin) implicates fork uncoupling as the initial checkpoint signalPhosphorylation of the Cep3 kinetochore protein by Dun1 kinase allows for centromere release, which is critical for collapsed fork repositioningDamage-inducible nuclear microtubules (DIMs) colocalize with the repeat locus and are required for relocation to the NPCEstablishes a new role for the DNA replication and DNA damage checkpoint response to trigger repositioning of collapsed forks within the nucleus.