Determinants of RPA megafoci localization to the nuclear periphery in response to replication stress

Area of Science:

• Cell Biology
• Molecular Biology
• Genetics

Background:

• Replication stress triggers the accumulation of single-stranded DNA (ssDNA), coated by replication protein A (RPA).
• This ssDNA-RPA complex signals the activation of the replication stress response pathway.
• Severe stress, such as loss of Mcm4, leads to large RPA foci at the nuclear periphery in Schizosaccharomyces pombe.

Purpose of the Study:

• To investigate the mechanisms underlying the formation and nuclear periphery translocation of large RPA foci under severe replication stress.
• To elucidate the role of resection, repair, and chromatin remodelers in this process.
• To understand how localized RPA accumulation impacts cell cycle regulation and the replication stress response.

Main Methods:

• Utilized a temperature-sensitive Schizosaccharomyces pombe degron mutant (mcm4-dg) to induce severe replication stress.
• Investigated the involvement of resection and repair pathways.
• Assessed the role of the Swr1/Ino80 chromatin remodeler complex.
• Analyzed RPA focus formation, localization, and recruitment of Cds1.
• Examined cell cycle progression, specifically Tos4 accumulation and MBF-mediated G1/S transition.

Main Results:

• Resection, repair processes, and the Swr1/Ino80 chromatin remodeler are crucial for the formation and peripheral relocalization of large RPA foci.
• Concentrated RPA accumulation at the nuclear periphery enhances Cds1 recruitment to chromatin.
• This aberrant response leads to a cell cycle phenotype lacking the typical MBF-mediated G1/S accumulation of Tos4.

Conclusions:

• Localized accumulation of RPA at the nuclear periphery represents a distinct replication stress response mechanism.
• This RPA-mediated response facilitates evasion of canonical cell cycle arrest.
• The findings highlight a novel pathway for managing severe replication stress and maintaining cell viability.