CDC45 is required in conjunction with CDC7/DBF4 to trigger the initiation of DNA replication.

The initiation of DNA replication in Saccharomyces cerevisiae requires the protein product of the CDC45 gene. We report that although Cdc45p is present at essentially constant levels throughout the cell cycle, it completes its initiation function in late G1, after START and prior to DNA synthesis. Shortly after mitosis, cells prepare for initiation by assembling prereplicative complexes at their replication origins. These complexes are then triggered at the onset of S phase to commence DNA replication. Cells defective for CDC45 are incapable of activating the complexes to initiate DNA replication. In addition, Cdc45p and Cdc7p/Dbf4p, a kinase implicated in the G1/S phase transition, are dependent on one another for function. These data indicate that CDC45 functions in late G1 phase in concert with CDC7/DBF4 to trigger initiation at replication origins after the assembly of the prereplicative complexes.

[1]  T. Kelly,et al.  DNA replication: Controlling initiation during the cell cycle , 1996, Current Biology.

[2]  C. F. Hardy Identification of Cdc45p, an essential factor required for DNA replication. , 1997, Gene.

[3]  J. Blow,et al.  The role of MCM/P1 proteins in the licensing of DNA replication. , 1996, Trends in biochemical sciences.

[4]  D. Engelke,et al.  Direct sequence and footprint analysis of yeast DNA by primer extension. , 1991, Methods in enzymology.

[5]  B. Futcher,et al.  The Cln3‐Cdc28 kinase complex of S. cerevisiae is regulated by proteolysis and phosphorylation. , 1992, The EMBO journal.

[6]  B. Tye,et al.  Mcm2 and Mcm3, two proteins important for ARS activity, are related in structure and function. , 1991, Genes & development.

[7]  A. Jackson,et al.  Cell cycle regulation of the yeast Cdc7 protein kinase by association with the Dbf4 protein , 1993, Molecular and cellular biology.

[8]  B. Tye,et al.  Cell cycle-regulated nuclear localization of MCM2 and MCM3, which are required for the initiation of DNA synthesis at chromosomal replication origins in yeast. , 1993, Genes & development.

[9]  F. Cross,et al.  CLB5: a novel B cyclin from budding yeast with a role in S phase. , 1992, Genes & development.

[10]  J. Li,et al.  Cdc6p establishes and maintains a state of replication competence during G1 phase. , 1997, Journal of cell science.

[11]  J. Rine,et al.  Origin recognition complex (ORC) in transcriptional silencing and DNA replication in S. cerevisiae. , 1993, Science.

[12]  B. Stillman,et al.  CDC45, a novel yeast gene that functions with the origin recognition complex and Mcm proteins in initiation of DNA replication , 1997, Molecular and cellular biology.

[13]  J. Rine,et al.  The origin recognition complex in silencing, cell cycle progression, and DNA replication. , 1995, Molecular biology of the cell.

[14]  S. Dalton,et al.  Cdc54 belongs to the Cdc46/Mcm3 family of proteins which are essential for initiation of eukaryotic DNA replication. , 1995, Gene.

[15]  T. Coleman,et al.  The Xenopus Cdc6 Protein Is Essential for the Initiation of a Single Round of DNA Replication in Cell-Free Extracts , 1996, Cell.

[16]  K Nasmyth,et al.  CLB5 and CLB6, a new pair of B cyclins involved in DNA replication in Saccharomyces cerevisiae. , 1993, Genes & development.

[17]  K. Nasmyth,et al.  Yeast origin recognition complex is involved in DNA replication and transcriptional silencing , 1993, Nature.

[18]  Kim Nasmyth,et al.  The B-type cyclin kinase inhibitor p40 SIC1 controls the G1 to S transition in S. cerevisiae , 1994, Cell.

[19]  J. Diffley,et al.  Interaction of Dbf4, the Cdc7 protein kinase regulatory subunit, with yeast replication origins in vivo. , 1994, Science.

[20]  D. Botstein,et al.  Subcellular localization of yeast CDC46 varies with the cell cycle. , 1990, Genes & development.

[21]  J. Diffley,et al.  Two steps in the assembly of complexes at yeast replication origins in vivo , 1994, Cell.

[22]  I. Todorov,et al.  The role of MCM proteins in the cell cycle control of genome duplication , 1996, BioEssays : news and reviews in molecular, cellular and developmental biology.

[23]  J. Rine,et al.  The origin recognition complex has essential functions in transcriptional silencing and chromosomal replication. , 1995, Genes & development.

[24]  I. Herskowitz,et al.  Isolation of ORC6, a component of the yeast origin recognition complex by a one-hybrid system. , 1993, Science.

[25]  J. Li,et al.  Eukaryotic DNA replication. , 1995, Current opinion in cell biology.

[26]  J. Diffley,et al.  Once and only once upon a time: specifying and regulating origins of DNA replication in eukaryotic cells. , 1996, Genes & development.

[27]  S. Dalton,et al.  Cell cycle-regulated nuclear import and export of Cdc47, a protein essential for initiation of DNA replication in budding yeast. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[28]  J. Diffley,et al.  Protein-DNA interactions at a yeast replication origin , 1992, Nature.

[29]  K. Nasmyth,et al.  An essential role for the Cdc6 protein in forming the pre-replicative complexes of budding yeast , 1996, Nature.

[30]  P. Pahl,et al.  mcm5/cdc46-bob1 bypasses the requirement for the S phase activator Cdc7p. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[31]  D. Botstein,et al.  A group of interacting yeast DNA replication genes. , 1991, Genes & Development.

[32]  S. Dalton,et al.  Cdc45p assembles into a complex with Cdc46p/Mcm5p, is required for minichromosome maintenance, and is essential for chromosomal DNA replication. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[33]  L. Hartwell Sequential function of gene products relative to DNA synthesis in the yeast cell cycle. , 1976, Journal of molecular biology.

[34]  R. Rothstein Targeting, disruption, replacement, and allele rescue: integrative DNA transformation in yeast. , 1991, Methods in enzymology.

[35]  S. Bell,et al.  Yeast origin recognition complex functions in transcription silencing and DNA replication. , 1993, Science.

[36]  A. Jackson,et al.  Cdc7 protein kinase for DNA metabolism comes of age , 1994, Molecular microbiology.

[37]  B. Tye The MCM2-3-5 proteins: are they replication licensing factors? , 1994, Trends in cell biology.

[38]  L. Drury,et al.  Cdc6p-dependent loading of Mcm proteins onto pre-replicative chromatin in budding yeast. , 1997, Proceedings of the National Academy of Sciences of the United States of America.