Exceptional origin activation revealed by comparative analysis in two laboratory yeast strains

We performed a comparative analysis of replication origin activation by genome-wide single-stranded DNA mapping in two yeast strains challenged by hydroxyurea, an inhibitor of the ribonucleotide reductase. We gained understanding of the impact on origin activation by three factors: S-phase checkpoint control, DNA sequence polymorphisms, and relative positioning of origin and transcription unit. Wild type W303 showed a significant reduction of fork progression accompanied by an elevated level of Rad53 phosphorylation as well as physical presence at origins compared to A364a. Moreover, a rad53K227A mutant in W303 activated more origins, accompanied by global reduction of ssDNA across all origins, compared to A364a. Sequence polymorphism in the consensus motifs of origins plays a minor role in determining strain-specific activity. Finally, we identified a new class of origins only active in checkpoint-proficient cells, which we named Rad53-dependent origins. Our study presents a comprehensive list of differentially used origins and provide new insights into the mechanisms of origin activation.

Automated summary

This study compared replication origin activation in two yeast strains under hydroxyurea treatment, revealing the roles of S-phase checkpoint control, DNA sequence polymorphisms, and origin-transcription unit positioning. Rad53 protein plays a significant role in activation of origins, particularly in checkpoint-proficient cells.