Cdc14 is released from the nucleolus under DNA damage and is required for DNA repair by homologous recombination

Abstract

Endogenous metabolic products, such as reactive oxygen species, and exogenous genotoxic stress, constantly assault the genetic material in the cell. In response to these sources of DNA damage, cells have developed a coordinated signalling network known as the DDR (DNA damage response), which coordinates cell cycle progression with DNA repair. Recently, multiple lines of evidences have suggested a complex role for several kinases in the regulation of the DDR (including the CDK), but little is known about the phosphatases that revert the effects of these kinases. The serine/threonine phosphatase Cdc14 was firstly identify in S. cerevisiae as an essential cell cycle phosphatase required for Cdk inactivation. This phosphatase is sequestered into the nucleolus during interphase due to its interaction with Net1. During anaphase, Net1 is phosphorylated and Cdc14 is released from the nucleolus to allow cells exit from mitosis. Cdc14 has predisposition to dephosphorylate Cdk targets, therefore is reasonable to think that could be a key candidate to counterbalance the effect imposed by the Cdk in the DDR. In favour of this hypothesis, Cdc14 is required for cell viability under different DNA damage conditions. To determine the role of the phosphatase in the DDR, we used two different recombinational repair pathways: SDSA (Synthensis dependent strand annealing) and dHJ (double Holliday junction repair). We observed that cells lacking Cdc14 activity presented defects in DNA repair in both SDSA and dHJ. Supporting the role of the phosphatase in DNA repair, we observed that Cdc14 was released from the nucleolus after induction of a single DSB or treatment with phleomycin. Accordingly, Net1 was phosphorylated during the execution of the DDR. Finally, by using mass spectrometry in a screen to identify targets of the phosphatase exclusively in DNA damage, we have detected several targets directly implicated in DNA damage repair. Altogether, we have placed Cdc14 at the DNA damage response context by collaborating in the repair throughout the modulation of the homologous recombination repair pathway, and providing new evidences about the role of this phosphatase in the repair of a DNA lesion.