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Related Experiment Videos

IS911-mediated intramolecular transposition is naturally temperature sensitive

L Haren1, M Bétermier, P Polard

  • 1Laboratoire de Microbiologie et Génétique Moléculaires, CNRS UPR9007,Toulouse, France.

Molecular Microbiology
|August 1, 1997
PubMed
Summary

The bacterial insertion sequence IS911 shows temperature-sensitive transposition. Its transposase, OrfAB, produces fewer DNA intermediates at higher temperatures, suggesting the transposase itself is heat-sensitive.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Microbiology

Background:

  • The bacterial insertion sequence IS911 is known to transpose within genomes.
  • Previous studies indicated that high levels of IS911 transposase (OrfAB) lead to specific DNA intermediate formations: a figure-eight form and an excised circular form.

Purpose of the Study:

  • To investigate the impact of temperature on the transposition activity of IS911.
  • To determine if the IS911 transposase (OrfAB) exhibits temperature-dependent behavior.

Main Methods:

  • In vivo and in vitro assays were used to monitor IS911 transposition.
  • Quantification of specific DNA intermediates (figure-eight and excised circular forms) at different temperatures (30°C, 37°C, and 42°C).
  • Analysis of two point mutants of OrfAB for temperature resistance.

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Main Results:

  • IS911 transposition and DNA intermediate formation were significantly reduced at 42°C compared to 37°C.
  • Lowering the temperature to 30°C increased the production of these intermediates.
  • A cell-free system and in vivo assays confirmed this temperature-activity profile for OrfAB.
  • Mutant OrfAB proteins showed partial resistance to temperature effects.

Conclusions:

  • The transposition of bacterial insertion sequence IS911 is highly sensitive to temperature.
  • The IS911 transposase (OrfAB) appears to have a temperature-sensitive property, affecting its activity at elevated temperatures.
  • These findings provide insights into the regulatory mechanisms of transposon mobility.