Rif1 Controls DNA Replication Timing in Yeast through the PP1 Phosphatase Glc7
Stefano Mattarocci,
Maksym Shyian,
Laure Lemmens,
Pascal Damay,
Dogus Murat Altintas,
Tianlai Shi,
Clinton R. Bartholomew,
Nicolas H. Thomä,
Christopher F.J. Hardy,
David Shore
Affiliations
Stefano Mattarocci
Department of Molecular Biology and Institute of Genetics and Genomics in Geneva, University of Geneva, 30 quai Ernest-Ansermet, 1211 Geneva, Switzerland
Maksym Shyian
Department of Molecular Biology and Institute of Genetics and Genomics in Geneva, University of Geneva, 30 quai Ernest-Ansermet, 1211 Geneva, Switzerland
Laure Lemmens
Department of Molecular Biology and Institute of Genetics and Genomics in Geneva, University of Geneva, 30 quai Ernest-Ansermet, 1211 Geneva, Switzerland
Pascal Damay
Department of Molecular Biology and Institute of Genetics and Genomics in Geneva, University of Geneva, 30 quai Ernest-Ansermet, 1211 Geneva, Switzerland
Dogus Murat Altintas
Department of Molecular Biology and Institute of Genetics and Genomics in Geneva, University of Geneva, 30 quai Ernest-Ansermet, 1211 Geneva, Switzerland
Tianlai Shi
Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, 4058 Basel, Switzerland
Clinton R. Bartholomew
Department of Cell and Developmental Biology, Vanderbilt University Medical Center, T-2212 Medical Center North, Nashville, TN 37232-2175, USA
Nicolas H. Thomä
Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, 4058 Basel, Switzerland
Christopher F.J. Hardy
Department of Cell and Developmental Biology, Vanderbilt University Medical Center, T-2212 Medical Center North, Nashville, TN 37232-2175, USA
David Shore
Department of Molecular Biology and Institute of Genetics and Genomics in Geneva, University of Geneva, 30 quai Ernest-Ansermet, 1211 Geneva, Switzerland
The Rif1 protein, originally identified as a telomere-binding factor in yeast, has recently been implicated in DNA replication control from yeast to metazoans. Here, we show that budding yeast Rif1 protein inhibits activation of prereplication complexes (pre-RCs). This inhibitory function requires two N-terminal motifs, RVxF and SILK, associated with recruitment of PP1 phosphatase (Glc7). In G1 phase, we show both that Glc7 interacts with Rif1 in an RVxF/SILK-dependent manner and that two proteins implicated in pre-RC activation, Mcm4 and Sld3, display increased Dbf4-dependent kinase (DDK) phosphorylation in rif1 mutants. Rif1 also interacts with Dbf4 in yeast two-hybrid assays, further implicating this protein in direct modulation of pre-RC activation through the DDK. Finally, we demonstrate Rif1 RVxF/SILK motif-dependent recruitment of Glc7 to telomeres and earlier replication of these regions in cells where the motifs are mutated. Our data thus link Rif1 to negative regulation of replication origin firing through recruitment of the Glc7 phosphatase.
