Site-specific recombination promotes plasmid amplification in yeast.

Area of Science:

• Molecular Biology
• Yeast Genetics
• Plasmids

Background:

• Stable yeast plasmids, like the 2-micron circle, possess long, inverted repeats that facilitate recombination.
• This recombination generates two inversion isomers present in equal cellular concentrations.
• The FLP recombinase, encoded by the 2-micron plasmid, catalyzes this site-specific inversion.

Purpose of the Study:

• To investigate the role of the 2-micron plasmid's site-specific recombination system in regulating plasmid copy number.
• To determine if FLP-mediated recombination influences yeast plasmid maintenance under nonselective conditions.

Main Methods:

• Analysis of 2-micron plasmid DNA in Saccharomyces cerevisiae.
• Investigating the impact of FLP recombinase activity on plasmid replication dynamics.
• Comparing plasmid copy numbers under varying conditions.

Main Results:

• The FLP-mediated site-specific recombination system of the 2-micron plasmid enhances its mean intracellular copy number in yeast.
• This increase in copy number is linked to a FLP-induced transient shift in replication from theta mode to double rolling circle mode.
• This mechanism appears to counteract copy number decreases caused by imperfect plasmid partitioning.

Conclusions:

• The 2-micron plasmid's recombination system is crucial for maintaining high and stable copy numbers.
• FLP-mediated replication mode switching serves as a copy number correction mechanism, ensuring plasmid stability.
• This study elucidates a novel strategy for plasmid maintenance in eukaryotic systems.