4E). is found at both the Cdc10 promoter and intragenic binding sites, Rep2 appears to be absent at the Cdc10 promoter binding sites but present at the intragenic sites. Time course ChIP-chip analysis demonstrates that Rep2 is temporally accumulated at the coding region of the MBF target genes, resembling the RNAP-II occupancies. Taken together, our results show that deconvolution analysis of Cdc10 occupancies refines the functional subset CIP1 of genomic binding sites. We propose that the MBF activator Rep2 plays a role in mediating the cell cycle-specific transcription through the recruitment of RNAP-II to the MBF-bound G1/S-phase genes. In eukaryotes, from budding yeast (12,50) and fission yeast (43,44,47) to mammalian cell line systems (13,56), cell cycle progression is accompanied by a periodic change in transcription levels of 400 to 1 1,000 genes. Transcriptional regulation plays a pivotal role in controlling the processes of cellular differentiation and division. Cell development fate is determined during G1at a restriction point known as START in yeast, which is marked by the coordinated induction of a subset of cell cycle-specific genes that promote cell mass/size increase and entry into S phase (27). InSaccharomyces cerevisiae, activation of the two heterodimeric transcription factor complexes SBF (Swi4/6-related cell cycle box [SCB]-bindingfactor) and MBF (MluI-like cell cycle box [MCB]-bindingfactor) is required for initiating a number of key events, such as budding, DNA synthesis, and spindle pole body duplication at the G1/S transition (32,33). Cell cycle-specific transcription of SWI4 is required for the cell cycle regulation of HO transcription (6) and possibly other Swi4/Swi6 target genes. Swi4/Swi6 binds to its genomic targets at the G1phase (26), prior to the activation by the CDK Cdc28/Cln3 (37) through the inactivation of the SBF repressor Whi5 (21). Thus, the SBF activity is coordinately controlled by both the CDK and the cell cycle transcription. Activated SBF induces the transcription of several G1cyclin-encoding genes, the HO-endonuclease gene, and genes required for cell wall biogenesis. On the other hand, the CDK-activated MBF induces the expression of the S-phase cyclin genes and genes required for DNA synthesis and is repressed by the Rb-like protein Nrm1 after S-phase entry (20). In animal Biotin Hydrazide cells, Rb is known to repress the G1-specific E2F-DP1 transcriptional complex (16,22), an analog of theS. cerevisiaeSBF/MBF complex. At the G1-S transition, CDK Cdk4/cyclin D inactivates Rb and thus relieves the repression of the E2F-DP1 complex (11). InSchizosaccharomyces pombe, the heterodimers Cdc10/Res1 (homolog of Swi6/Mbp1) and Cdc10/Res2 (homolog of Swi6/Swi4) play essential roles in the G1/S transition (4,10,36,39,40,42,51,54,60). It is proposed that Cdc10/Res1 and Cdc10/Res2 constitute the core MBF complex and coordinately bind to the MCB-like motif (55). Cdc10 has been shown to bind the promoter ofcdc18constitutively (58). Given that Cdc10 forms complexes with Res1 and Res2 throughout the cell cycle (55), it is conceivable that the core MBF complex Res1-Cdc10-Res2 is bound at the promoter of the target genes throughout the cell cycle. However, activation of MBF at the G1phase is complex. It has been shown that MBF is activated by Cdc2 in the G1phase (17,18,46). On the other hand, the MBF activity is also regulated by the CDK Pef1-Pas1 (52). It is therefore likely that MBF activity is regulated by multiple signaling pathways inS. pombe. After G1/S-phase transcription, the MBF activity is likely to be repressed by a number of negative regulators, Cig2, Nrm1, and Yox1 (1,2,20). The activation of MBF requires the activator Rep2 (42). The constitutively active allelecdc10-C4is also known to be dependent on the Biotin Hydrazide function of Rep2 (55), suggesting that Rep2 is absolutely essential for MBF activity. Although the role of Rep2 in the activation of MBF is unclear, we have previously shown that Rep2 protein levels oscillated during cell cycle, with a peak level at G1/S phase, consistent with its role in the activation of MBF transcriptional activity (14). Over 200 genes targeted Biotin Hydrazide by SBF/MBF in theS. cerevisiaegenome have been identified (28). Many SBF/MBF target genes are found to contain no apparent SCB/MCB promoter motif, suggesting that SBF/MBF can bind the SCB/MCB-less sequences, possibly through the interaction with other transcription factors. Notably, transcription of many SBF/MBF target genes is not cell cycle regulated, suggesting that SBF/MBF plays a role in regulation of both periodic and constitutive transcription. InS. pombe, 80 binding targets for Cdc10 and Yox1, half of which are not periodically transcribed, have been identified (1). This raises a.