G1-specific transcriptional activation by Cln3/CDK initiates the budding yeast cell cycle. To identify targets of Cln3/CDK, we analyzed the SBF and MBF transcription factor complexes by multidimensional protein interaction technology (MudPIT). Whi5 was identified as a stably bound component of SBF but not MBF. Inactivation of Whi5 leads to premature expression of G1-specific genes and budding, whereas overexpression retards those processes. Whi5 inactivation bypasses the requirement for Cln3 both for transcriptional activation and cell cycle initiation. Whi5 associates with G1-specific promoters via SBF during early G1 phase, then dissociates coincident with transcriptional activation. Dissociation of Whi5 is promoted by Cln3 in vivo. Cln/CDK phosphorylation of Whi5 in vitro promotes its dissociation from SBF complexes. Mutation of putative CDK phosphorylation sites, at least five of which are phosphorylated in vivo, strongly reduces SBF-dependent transcription and delays cell cycle initiation. Like mammalian Rb, Whi5 is a G1-specific transcriptional repressor antagonized by CDK.

The Mre11-Rad50-Nbs1 (MRN) complex is a primary sensor of DNA double-strand breaks (DSBs). Upon recruitment to DSBs, it plays a critical role in catalyzing 5' --> 3' single-strand resection that is required for repair by homologous recombination (HR). Unknown mechanisms repress HR in G1 phase of the cell cycle during which nonhomologous end-joining (NHEJ) is the favored mode of DSB repair. Here we describe fission yeast Ctp1, so-named because it shares conserved domains with the mammalian tumor suppressor CtIP. Ctp1 is recruited to DSBs where it is essential for repair by HR. Ctp1 is required for efficient formation of RPA-coated single-strand DNA adjacent to DSBs, indicating that it functions with the MRN complex in 5' --> 3' resection. Transcription of ctp1(+) is periodic during the cell cycle, with the onset of its expression coinciding with the start of DNA replication. These data suggest that regulation of Ctp1 underlies cell-cycle control of HR.

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