Abstract
Fork stabilization at DNA impediments is key to maintaining replication fork integrity and preventing chromosome breaks. Mrc1 and Tof1 are two known stabilizers that travel with the replication fork. In addition to a structural role, Mrc1 has a DNA damage checkpoint function. Using a yeast model system, we analyzed the role of Mrc1 and Tof1 at expanded CAG repeats of medium and long lengths, which are known to stall replication forks and cause trinucleotide expansion diseases such as Huntington's disease and myotonic dystrophy. We demonstrate that the fork stabilizer but not the checkpoint activation function of Mrc1 is key for preventing DNA breakage and death of cells containing expanded CAG tracts. In contrast, both Mrc1 functions are important in preventing repeat length instability. Mrc1 has a general fork protector role that is evident at forks traversing both repetitive and non-repetitive DNA, though it becomes crucial at long CAG repeat lengths. In contrast, the role of Tof1 in preventing fork breakage is specific to long CAG tracts of 85 or more repeats. Our results indicate that long CAG repeats have a particular need for Tof1 and highlight the importance of fork stabilizers in maintaining fork integrity during replication of structure-forming repeats.
Highlights
DNA replication is a robust process that allows the transmission of the genetic information to a daughter cell with a high level of fidelity
CAG fragility was measured by a genetic assay that detects chromosome endloss resulting from chromosome breakage at or near the repeat tract; a telomere seed sequence proximal to the repeat facilitates recovery of broken chromosomes, which results in loss of the distal URA3 gene and 5-FOA resistant colonies (Supplementary Figure S1)
Comparison of the mrc1, mrc1AQ and rad53-21 mutants revealed that the DNA damage checkpoint regulated by Rad53 is important for promoting cell division and preventing chromosome fragility
Summary
DNA replication is a robust process that allows the transmission of the genetic information to a daughter cell with a high level of fidelity. Replication faces numerous impediments that perturb its progression and can lead to a replication fork stall. These impediments can be a tightly bound protein, damaged or cross-linked nucleotides, or DNA structures [1,2,3]. In order to achieve fork restart and completion of replication, the stalled fork needs to be stabilized. Replicative stress is the hallmark of cells with activated oncogenes, and is one cause of the genome instability that occurs in early stages of tumorigenesis [4,5,6,7]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
