A role for the Cdc7 kinase regulatory subunit Dbf4p in the formation of initiation-competent origins of replication (original) (raw)

  1. Philippe Pasero,
  2. Bernard P. Duncker,
  3. Etienne Schwob, and
  4. Susan M. Gasser
  5. Swiss Institute for Experimental Cancer Research (ISREC), CH-1066 Epalinges/Lausanne, Switzerland; Institute of Molecular Genetics, UMR 5535, Centre National de la Recherche Scientifique, F-34033 Montpellier, France

Abstract

Using a reconstituted DNA replication assay from yeast, we demonstrate that two kinase complexes are essential for the promotion of replication in vitro. An active Clb/Cdc28 kinase complex, or its vertebrate equivalent, is required in trans to stimulate initiation in G1-phase nuclei, whereas the Dbf4/Cdc7 kinase complex must be provided by the template nuclei themselves. The regulatory subunit of Cdc7p, Dbf4p, accumulates during late G1 phase, becomes chromatin associated prior to Clb/Cdc28 activation, and assumes a punctate pattern of localization that is similar to, and dependent on, the origin recognition complex (ORC). The association of Dbf4p with a detergent-insoluble chromatin fraction in G1-phase nuclei requires ORC but not Cdc6p or Clb/Cdc28 kinase activity, and correlates with competence for initiation. We propose a model in which Dbf4p targets Cdc7p to the prereplication complex prior to the G1/S transition, by a pathway parallel to, but independent of, the Cdc6p-dependent recruitment of MCMs.

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