EXO1-dependent single-stranded DNA at telomeres activates subsets of DNA damage and spindle checkpoint pathways in budding yeast yku70Δ mutants

Abstract

We have examined the role of checkpoint pathways in responding to ayku70Δ defect in budding yeast. We show thatCHK1, MEC1, and RAD9 checkpoint genes are required for efficient cell cycle arrest of yku70Δ mutants cultured at 37°C, whereas RAD17,RAD24, MEC3, DDC1, and DUN1play insignificant roles. We establish that cell cycle arrest ofyku70Δ mutants is associated with increasing levels of single-stranded DNA in subtelomeric Y‘ regions, and find that the mismatch repair-associated EXO1 gene is required for both ssDNA generation and cell cycle arrest of yku70Δ mutants. In contrast, MRE11 is not required for ssDNA generation. The behavior of yku70Δ exo1Δ double mutants strongly indicates that ssDNA is an important component of the arrest signal inyku70Δ mutants and demonstrates a link between damaged telomeres and mismatch repair-associated exonucleases. This link is confirmed by our demonstration that EXO1 also plays a role in ssDNA generation in cdc13-1 mutants. We have also found that the MAD2 but not the BUB2 spindle checkpoint gene is required for efficient arrest of yku70Δ mutants. Therefore, subsets of both DNA-damage and spindle checkpoint pathways cooperate to regulate cell division of yku70Δ mutants.