title: DNA polymerase switching: I. Replication factor C displaces DNA polymerase alpha prior to PCNA loading.
abstract: An important not yet fully understood event in DNA replication is the DNA polymerase (pol) switch from pol alpha to pol delta. Indirect evidence suggested that the clamp loader replication factor C (RF-C) plays an important role, since a replication competent protein complex containing pol alpha, pol delta and RF-C could perform pol switching in the presence of proliferating cell nuclear antigen (PCNA). By using purified pol alpha/primase, pol delta, RF-C, PCNA and RP-A we show that: (i) RF-C can inhibit pol alpha in the presence of ATP prior to PCNA loading, (ii) RF-C decreases the affinity of pol alpha for the 3'OH primer ends, (iii) the inhibition of pol alpha by RF-C is released upon PCNA loading, (iv) ATP hydrolysis is required for PCNA loading and subsequent release of inhibition of pol alpha, (v) under these conditions a switching from pol alpha/primase to pol delta is evident. Thus, RF-C appears to be critical for the pol alpha to pol delta switching. Based on these results, a model is proposed in which RF-C induces the pol switching by sequestering the 3'-OH end from pol alpha and subsequently recruiting PCNA to DNA.
authors.affiliation: Istituto di Genetica Biochimica ed Evoluzionistica IGBE-CNR, Pavia, I-27100, Italy.
authors.forename: G
authors.initials: G
authors.lastname: Maga
authors.forename: M
authors.initials: M
authors.lastname: Stucki
authors.forename: S
authors.initials: S
authors.lastname: Spadari
authors.forename: U
authors.initials: U
authors.lastname: Hübscher
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002417:Cattle; D003600:Cytosol / Q000201:enzymology; D004256:DNA Polymerase I / Q000378:metabolism*; D004258:DNA Polymerase III / Q000378:metabolism*; D019915:DNA Primase / Q000378:metabolism; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells; D018398:Homeodomain Proteins; D006801:Humans; D007700:Kinetics; D015778:Minor Histocompatibility Antigens; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2; D051818:Replication Protein C; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D013539:Simian virus 40 / Q000235:genetics / Q000502:physiology; D013379:Substrate Specificity; D013950:Thymus Gland / Q000201:enzymology; D014779:Virus Replication
chemical_list: C096816:BCL2-related protein A1; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; C096063:MATA1 protein, S cerevisiae; D015778:Minor Histocompatibility Antigens; D018809:Proliferating Cell Nuclear Antigen; D019253:Proto-Oncogene Proteins c-bcl-2; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D000255:Adenosine Triphosphate; D019915:DNA Primase; D004256:DNA Polymerase I; D004258:DNA Polymerase III; D051818:Replication Protein C