Abstract
Despite its evolutionarily conserved function in controlling DNA replication, the chromosomal binding sites of the budding yeast Rif1 protein are not well understood. Here, we analyse genome‐wide binding of budding yeast Rif1 by chromatin immunoprecipitation, during G1 phase and in S phase with replication progressing normally or blocked by hydroxyurea. Rif1 associates strongly with telomeres through interaction with Rap1. By comparing genomic binding of wild‐type Rif1 and truncated Rif1 lacking the Rap1‐interaction domain, we identify hundreds of Rap1‐dependent and Rap1‐independent chromosome interaction sites. Rif1 binds to centromeres, highly transcribed genes and replication origins in a Rap1‐independent manner, associating with both early and late‐initiating origins. Interestingly, Rif1 also binds around activated origins when replication progression is blocked by hydroxyurea, suggesting association with blocked forks. Using nascent DNA labelling and DNA combing techniques, we find that in cells treated with hydroxyurea, yeast Rif1 stabilises recently synthesised DNA. Our results indicate that, in addition to controlling DNA replication initiation, budding yeast Rif1 plays an ongoing role after initiation and controls events at blocked replication forks.
Original language | English |
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Article number | e46222 |
Number of pages | 16 |
Journal | EMBO reports |
Volume | 19 |
Issue number | 9 |
Early online date | 13 Aug 2018 |
DOIs | |
Publication status | Published - 1 Sept 2018 |
Bibliographical note
We thank Javier Garzon and Vamsi Krishna Gali for discussion and advice on methods, and Takashi Kubota for helpful comments on the manuscript. This work was supported by Cancer Research UK Programme Award A19059 to ADD and SH. KS was supported by Grant‐in‐Aid for Scientific Research on Priority Areas (15H05970 and 15K21761) from Ministry of Education, Culture, Sports, Science and Technology, Japan.Funding
Cancer Research UK (CRUK) A19059
Ministry of Education, Culture, Sports, Science and Technology (MEXT) 15H0597015K21761
Corrigendum to Budding yeast Rif1 binds to replication origins and protects DNA at blocked replication forks (EMBO reports, (2018), 19, 9, (e46222), 10.15252/embr.201846222)
It came to the authors’ attention that in the original paper, an incorrect conversion factor was used for the pixel to micrometre conversion when analysing microscope images of combed fibres. The conversion factor used was 7.2 pixels = 1 µm instead of 13.9 pixels = 1 µm.
Data Availability Statement
Data availabilityChIP‐Seq data and corresponding input data were submitted to ArrayExpress under accession number E‐MTAB‐6736.
Keywords
- Rif1
- DNA replication origin
- ChIP-Seq
- Nascent DNA
- Centromere
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Anne Donaldson
- School of Medicine, Medical Sciences & Nutrition, Molecular and Cellular Function
- School of Medicine, Medical Sciences & Nutrition, Aberdeen Cancer Centre
- School of Medicine, Medical Sciences & Nutrition, Medical Sciences - Personal Chair
- School of Medicine, Medical Sciences & Nutrition, Institute of Medical Sciences
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Centre for Genome-Enabled Biology and Medicine
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