Polymerase Epsilon Is Required To Maintain Replicative Senescence

Abstract

Replicative senescence is a permanent cell cycle arrest in response to extensive telomere shortening. To understand the mechanisms behind a permanent arrest, we screened for factors affecting replicative senes-cence in budding yeast lacking telomere elongation pathways. Intriguingly, we found that DNA polymerase epsilon (Pol ␧) acts synergistically with Exo1 nuclease to maintain replicative senescence. In contrast, the Pol ␧-associated checkpoint and replication protein Mrc1 facilitates escape from senescence. To understand this paradox, in which DNA-synthesizing factors cooperate with DNA-degrading factors to maintain arrest, whereas a checkpoint protein opposes arrest, we analyzed the dynamics of double-and single-stranded DNA (ssDNA) at chromosome ends during senescence. We found evidence for cycles of DNA resection, followed by resynthesis. We propose that resection of the shortest telomere, activating a Rad24 Rad17 -dependent checkpoint pathway, alternates in time with an Mrc1-regulated Pol ␧ resynthesis of a short, double-stranded chromosome end, which in turn activates a Rad9 53BP1 -dependent checkpoint pathway. Therefore, instead of one type of DNA damage, different types (ssDNA and a double-strand break-like structure) alternate in a " vicious circle, " each activating a different checkpoint sensor. Every time resection and resynthesis switches, a fresh signal initiates, thus preventing checkpoint adaptation and ensuring the permanent character of senescence.