Investigating the role of Rts1 in DNA replication initiation

Wellcome Open Research

article

Ana B.A. Wallis¹  Conrad A. Nieduszynski¹ 

[1] Sir William Dunn School of Pathology, University of Oxford

关键词: DNA Replication;    Phosphatases;    Rts1;    Rif1;    Orc2;   

DOI  :  10.12688/wellcomeopenres.13884.1

学科分类：内科医学

来源: Wellcome

【 摘 要 】

Background: Understanding DNA replication initiation is essential to understand the mis-regulation of replication seen in cancer and other human disorders. DNA replication initiates from DNA replication origins. In eukaryotes, replication is dependent on cell cycle kinases which function during S phase. Dbf4-dependent kinase (DDK) and cyclin-dependent kinase (CDK) act to phosphorylate the DNA helicase (composed of mini chromosome maintenance proteins: Mcm2-7) and firing factors to activate replication origins. It has recently been found that Rif1 can oppose DDK phosphorylation. Rif1 can recruit protein phosphatase 1 (PP1) to dephosphorylate MCM and restricts origin firing. In this study, we investigate a potential role for another phosphatase, protein phosphatase 2A (PP2A), in regulating DNA replication initiation. The PP2A regulatory subunit Rts1 was previously identified in a large-scale genomic screen to have a genetic interaction withORC2 (a DNA replication licensing factor). Deletion ofRTS1 synthetically rescued the temperature-sensitive (ts-) phenotype ofORC2 mutants.Methods: We deletedRTS1 in multiple ts-replication factorSaccharomyces cerevisiae strains, includingORC2.  Dilution series assays were carried out to compare qualitatively the growth of double mutant∆rts1 ts-replication factor strains relative to the respective single mutant strains.  Results: No synthetic rescue of temperature-sensitivity was observed. Instead we found an additive phenotype, indicating gene products function in separate biological processes. These findings are in agreement with a recent genomic screen which found thatRTS1 deletion in several ts-replication factor strains led to increased temperature-sensitivity.Conclusions: We find no evidence that Rts1 is involved in the dephosphorylation of DNA replication initiation factors.

【 授权许可】

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