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Bioinformatical dissection of fission yeast DNA replication origins.

ABSTRACT: Replication origins in eukaryotes form a base for assembly of the pre-replication complex (pre-RC), thereby serving as an initiation site of DNA replication. Characteristics of replication origin vary among species. In fission yeast Schizosaccharomyces pombe, DNA of high AT content is a distinct feature of replication origins; however, it remains to be understood what the general molecular architecture of fission yeast origin is. Here, we performed ChIP-seq mapping of Orc4 and Mcm2, two representative components of the pre-RC, and described the characteristics of their binding sites. The analysis revealed that fission yeast efficient origins are associated with two similar but independent features: a ?15 bp-long motif with stretches of As and an AT-rich region of a few hundred bp. The A-rich motif was correlated with chromosomal binding of Orc, a DNA-binding component in the pre-RC, whereas the AT-rich region was associated with efficient binding of the DNA replicative helicase Mcm. These two features, in combination with the third feature, a transcription-poor region of approximately 1 kb, enabled to distinguish efficient replication origins from the rest of chromosome arms with high accuracy. This study, hence, provides a model that describes how multiple functional elements specify DNA replication origins in fission yeast genome.

SUBMITTER: Masuda K

PROVIDER: S-EPMC7574548 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Bioinformatical dissection of fission yeast DNA replication origins.

Masuda Koji K Renard-Guillet Claire C Shirahige Katsuhiko K Sutani Takashi T

Open biology 20200722 7

Replication origins in eukaryotes form a base for assembly of the pre-replication complex (pre-RC), thereby serving as an initiation site of DNA replication. Characteristics of replication origin vary among species. In fission yeast <i>Schizosaccharomyces pombe</i>, DNA of high AT content is a distinct feature of replication origins; however, it remains to be understood what the general molecular architecture of fission yeast origin is. Here, we performed ChIP-seq mapping of Orc4 and Mcm2, two r ...[more]

PMID: 32692956

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Project description:BackgroundIn budding yeast, the replication checkpoint slows progress through S phase by inhibiting replication origin firing. In mammals, the replication checkpoint inhibits both origin firing and replication fork movement. To find out which strategy is employed in the fission yeast, Schizosaccharomyces pombe, we used microarrays to investigate the use of origins by wild-type and checkpoint-mutant strains in the presence of hydroxyurea (HU), which limits the pool of deoxyribonucleoside triphosphates (dNTPs) and activates the replication checkpoint. The checkpoint-mutant cells carried deletions either of rad3 (which encodes the fission yeast homologue of ATR) or cds1 (which encodes the fission yeast homologue of Chk2).ResultsOur microarray results proved to be largely consistent with those independently obtained and recently published by three other laboratories. However, we were able to reconcile differences between the previous studies regarding the extent to which fission yeast replication origins are affected by the replication checkpoint. We found (consistent with the three previous studies after appropriate interpretation) that, in surprising contrast to budding yeast, most fission yeast origins, including both early- and late-firing origins, are not significantly affected by checkpoint mutations during replication in the presence of HU. A few origins (approximately 3%) behaved like those in budding yeast: they replicated earlier in the checkpoint mutants than in wild type. These were located primarily in the heterochromatic subtelomeric regions of chromosomes 1 and 2. Indeed, the subtelomeric regions defined by the strongest checkpoint restraint correspond precisely to previously mapped subtelomeric heterochromatin. This observation implies that subtelomeric heterochromatin in fission yeast differs from heterochromatin at centromeres, in the mating type region, and in ribosomal DNA, since these regions replicated at least as efficiently in wild-type cells as in checkpoint-mutant cells.ConclusionThe fact that approximately 97% of fission yeast replication origins - both early and late - are not significantly affected by replication checkpoint mutations in HU-treated cells suggests that (i) most late-firing origins are restrained from firing in HU-treated cells by at least one checkpoint-independent mechanism, and (ii) checkpoint-dependent slowing of S phase in fission yeast when DNA is damaged may be accomplished primarily by the slowing of replication forks.

Dataset Information

Bioinformatical dissection of fission yeast DNA replication origins.

Publications

Bioinformatical dissection of fission yeast DNA replication origins.

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