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Related Experiment Videos

The replication fork barrier site forms a unique structure with Fob1p and inhibits the replication fork.

Takehiko Kobayashi1

  • 1National Institute for Basic Biology, 38 Nishigonaka, Myodaijicho, Okazaki 444-8585, Japan. koba@nibb.ac.jp

Molecular and Cellular Biology
|December 4, 2003
PubMed
Summary

The FOB1 protein directly binds to the replication fork barrier (RFB) site in yeast ribosomal DNA (rDNA). This interaction, mediated by a zinc finger motif, is crucial for blocking replication forks and driving rDNA recombination.

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Area of Science:

  • Molecular biology
  • Yeast genetics
  • DNA replication and recombination

Background:

  • The replication fork barrier (RFB) site in Saccharomyces cerevisiae rDNA is essential for regulating DNA replication.
  • The FOB1 gene product, Fob1p, is known to be required for RFB activity and various rDNA recombination events.
  • The precise molecular mechanism by which Fob1p functions in replication fork blocking and rDNA recombination remains unclear.

Purpose of the Study:

  • To elucidate the molecular mechanism of Fob1p's role in replication fork blocking and rDNA recombination.
  • To determine if Fob1p directly interacts with the RFB DNA sequence.
  • To identify the functional domains of Fob1p responsible for these activities.

Main Methods:

  • Chromatin immunoprecipitation (ChIP) assays

Related Experiment Videos

  • Gel shift assays
  • DNA footprinting analysis
  • Atomic force microscopy (AFM)

    • Main Results:

    • Fob1p directly binds to the RFB DNA sequence.
    • Fob1p interacts with two distinct sites within the RFB.
    • A predicted zinc finger motif in Fob1p is essential for RFB binding, replication fork blocking, and rDNA recombination.
    • The RFB DNA appears to wrap around Fob1p, suggesting a structural role in rDNA regulation.

    Conclusions:

    • Fob1p directly binds to the yeast rDNA RFB site.
    • The zinc finger motif of Fob1p is critical for its DNA-binding and functional activities.
    • The interaction between Fob1p and the RFB, potentially involving DNA wrapping, is key to regulating replication fork progression and rDNA recombination.