MCB Accepts, published online ahead of print on 2 November 2009

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Mol. Cell. Biol. doi:10.1128/MCB.01000-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Dephosphorylation of H2A by Glc7/PP1 promotes recovery from inhibition of DNA replication

Marco Bazzi, Davide Mantiero, Camilla Trovesi, Giovanna Lucchini, and Maria Pia Longhese^*

Dipartimento di Biotecnologie e Bioscienze, Università degli Studi di Milano-Bicocca, 20126 Milan, Italy

^* To whom correspondence should be addressed. Email: mariapia.longhese{at}unimib.it.

Replication fork stalling caused by deoxynucleotide depletion triggers Rad53 phosphorylation and subsequent checkpoint activation, which in turn plays a crucial role in maintaining functional DNA replication forks. How cells regulate checkpoint deactivation after inhibition of DNA replication is poorly understood. Here, we show that the budding yeast protein phosphatase Glc7/PP1 promotes disappearance of phosphorylated Rad53 and recovery from replication fork stalling caused by the dNTP synthesis inhibitor hydroxyurea (HU). Glc7 is also required for recovery from a DSB-induced checkpoint, while it is dispensable for checkpoint inactivation during MMS exposure, which instead requires the protein phosphatases Pph3, Ptc2 and Ptc3. Furthermore, Glc7 counteracts in vivo histone H2A phosphorylation on serine 129 (H2A) and dephosphorylates H2A in vitro. Finally, the replication recovery defects of HU-treated glc7 mutants are partially rescued by Rad53 inactivation or lack of H2A formation, and the latter also counteracts hyperphosphorylated Rad53 accumulation. We therefore propose that Glc7 activity promotes recovery from replication fork stalling caused by dNTP depletion, and that H2A dephosphorylation is a critical Glc7 function in this process.