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Related Experiment Videos

Antiparallel plasmid-plasmid pairing may control P1 plasmid replication.

A L Abeles1, S J Austin

  • 1Laboratory of Chromosome Biology, National Cancer Institute-Frederick Cancer Research and Development Center, MD 21702-1201.

Proceedings of the National Academy of Sciences of the United States of America
|October 15, 1991
PubMed
Summary
This summary is machine-generated.

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The P1 plasmid

Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • The P1 plasmid replicon maintains a strict copy number of one to two per cell.
  • Replication control is mediated by the copy-control locus incA, featuring RepA protein binding sites.
  • The replication origin also contains RepA binding sites crucial for initiating replication.

Purpose of the Study:

  • To investigate the mechanism by which the incA locus controls P1 plasmid replication.
  • To determine if incA sequences inhibit origin function in a trans-acting manner.
  • To elucidate the role of plasmid-plasmid interactions in replication control.

Main Methods:

  • Utilized an in vitro replication system with Escherichia coli extract, P1 origin template, and purified RepA protein.

Related Experiment Videos

  • Assessed the effect of supercoiled DNA circles containing the incA locus on origin function.
  • Varied the ratio of origin to incA sequences and RepA protein concentration.

    • Main Results:

    • Supercoiled DNA circles with the incA locus blocked P1 origin function in trans.
    • Complete shutdown of replication occurred at a 1:1 ratio of origin to incA sequences.
    • Excess RepA protein did not restore replication, indicating the mechanism is not simple protein titration.
    • incA sequences appear to inhibit replication through direct contact, favoring plasmid-plasmid pairing over cis-looping.

    Conclusions:

    • The incA locus controls P1 plasmid replication by directly interacting with the replication origin in a trans-acting manner.
    • Replication is inhibited through a plasmid-plasmid pairing mechanism where daughter plasmids' origins contact the incA locus of their partners.
    • This mechanism ensures stringent control over plasmid copy number per cell.