A second proliferating cell nuclear antigen loader complex, Ctf18-replication factor C, stimulates DNA polymerase η activity

Yasushi Shiomi, Chikahide Masutani, Fumio Hanaoka, Hiroshi Kimura, Toshiki Tsurimoto

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

Replication factor C (RFC) loads the clamp protein PCNA ontoDNAstructures. Ctf18-RFC, which consists of the chromosome cohesion factors Ctf18, Dcc1, and Ctf8 and four small RFC subunits, functions as a second proliferating cell nuclear antigen (PCNA) loader. To identify potential targets of Ctf18-RFC, human cell extracts were assayed for DNA polymerase activity specifically stimulated by Ctf18-RFC in conjunction with PCNA. After several chromatography steps, an activity stimulated by Ctf18-RFC but not by RFC was identified. Liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis revealed the presence of two DNA polymerases, η and λ, in the most purified fraction, but experiments with purified recombinant proteins demonstrated that only polymerase (pol) η was responsible for activity. Ctf18-RFC alone stimulated pol η, and the addition of PCNA cooperatively increased stimulation. Furthermore, Ctf18-RFC interacted physically with pol η, as indicated by co-precipitation in human cells. We propose that this novel loader-DNA polymerase interaction allows DNA replication forks to overcome interference by various template structures, including damaged DNA and DNA-protein complexes that maintain chromosome cohesion.

Original languageEnglish
Pages (from-to)20906-20914
Number of pages9
JournalJournal of Biological Chemistry
Volume282
Issue number29
DOIs
Publication statusPublished - Jul 20 2007

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Replication Protein C
Loaders
Proliferating Cell Nuclear Antigen
DNA-Directed DNA Polymerase
Chromosomes
DNA
Cells
Liquid chromatography
Clamping devices
Coprecipitation
Tandem Mass Spectrometry
Chromatography
Cell Extracts
DNA Replication
Recombinant Proteins
Liquid Chromatography
Mass spectrometry
Proteins

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

A second proliferating cell nuclear antigen loader complex, Ctf18-replication factor C, stimulates DNA polymerase η activity. / Shiomi, Yasushi; Masutani, Chikahide; Hanaoka, Fumio; Kimura, Hiroshi; Tsurimoto, Toshiki.

In: Journal of Biological Chemistry, Vol. 282, No. 29, 20.07.2007, p. 20906-20914.

Research output: Contribution to journalArticle

Shiomi, Yasushi ; Masutani, Chikahide ; Hanaoka, Fumio ; Kimura, Hiroshi ; Tsurimoto, Toshiki. / A second proliferating cell nuclear antigen loader complex, Ctf18-replication factor C, stimulates DNA polymerase η activity. In: Journal of Biological Chemistry. 2007 ; Vol. 282, No. 29. pp. 20906-20914.
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