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.