Spermidine biases the resolution of Holliday junctions by phage lambda integrase

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Holliday junctions are critical intermediates in DNA repair and recombination.
• Junction isomerization dictates recombination directionality, potentially influenced by DNA sequence.
• Understanding factors controlling isomerization is key to DNA metabolism pathways.

Purpose of the Study:

• To investigate the role of polycations in biasing strand cleavage during Holliday junction resolution.
• To determine if polycations can counteract sequence-specific biases in recombination.
• To elucidate the mechanism by which spermidine influences junction isomerization and resolution.

Main Methods:

• Studied wild-type phage lambda excision junctions.
• Assessed the effect of spermidine on junction isomerization and resolution rates.
• Examined spermidine's impact on both supercoiled and linear recombination intermediates.
• Investigated the requirement for accessory factors and their interaction with spermidine and DNA.

Main Results:

• Polycations, particularly spermidine, are major determinants of strand cleavage bias in Holliday junctions when sequence preference is absent.
• Spermidine significantly increases the rate of junction resolution and counteracts sequence-imposed biases.
• The spermidine effect is independent of DNA topology (supercoiled vs. linear) and requires accessory factors.
• Spermidine induces repositioning of Int protein domains, likely via DNA-spermidine-protein interactions or altered DNA conformation.

Conclusions:

• Spermidine, with accessory factors, promotes the formation of a specific Holliday junction isomer.
• This spermidine-induced isomerization likely activates Int active sites for efficient Holliday junction resolution in phage lambda recombination.