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Related Experiment Videos

Separation of origin recognition complex functions by cross-species complementation

A E Ehrenhofer-Murray1, M Gossen, D T Pak

  • 1Department of Molecular and Cell Biology, University of California, Berkeley 94720, USA.

Science (New York, N.Y.)
|December 8, 1995
PubMed
Summary
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The origin recognition complex (ORC) initiates DNA replication and regulates transcriptional silencing in yeast. A Drosophila melanogaster Orc2 protein variant could restore silencing but not replication, separating these ORC functions.

Area of Science:

  • Molecular Biology
  • Epigenetics
  • Yeast Genetics

Background:

  • Transcriptional silencing at the HMRa locus in Saccharomyces cerevisiae depends on the origin recognition complex (ORC).
  • The ORC is crucial for initiating DNA replication in yeast.
  • The orc2-1 yeast strain exhibits defects in both replication and silencing.

Purpose of the Study:

  • To investigate the distinct roles of the origin recognition complex (ORC) in transcriptional silencing and DNA replication.
  • To determine if the replication and silencing functions of ORC are separable.
  • To characterize the epigenetic nature of ORC-mediated silencing.

Main Methods:

  • Complementation analysis using Drosophila melanogaster Orc2 cDNA in a yeast orc2-1 mutant strain.
  • Mapping of the orc2-1 mutation to identify conserved regions between yeast and Drosophila ORC proteins.

Related Experiment Videos

  • Assessment of epigenetic propagation of the silent state during mitosis and its erasure during meiosis.

    • Main Results:

    • Expression of Drosophila Orc2 complemented the silencing defect but not the replication defect in the yeast orc2-1 strain.
    • This indicates that the replication and silencing functions of ORC are separable.
    • The silencing mediated by Drosophila Orc2 was epigenetic, stable through mitosis, but erased during meiosis.

    Conclusions:

    • The origin recognition complex (ORC) possesses distinct, separable functions in DNA replication and transcriptional silencing.
    • The Drosophila Orc2 protein can mediate epigenetic transcriptional silencing in yeast.
    • ORC-mediated silencing is mitotically stable but meiotically reversible.