title: A novel checkpoint and RPA inhibitory pathway regulated by Rif1.
abstract: Cells accumulate single-stranded DNA (ssDNA) when telomere capping, DNA replication, or DNA repair is impeded. This accumulation leads to cell cycle arrest through activating the DNA-damage checkpoints involved in cancer protection. Hence, ssDNA accumulation could be an anti-cancer mechanism. However, ssDNA has to accumulate above a certain threshold to activate checkpoints. What determines this checkpoint-activation threshold is an important, yet unanswered question. Here we identify Rif1 (Rap1-Interacting Factor 1) as a threshold-setter. Following telomere uncapping, we show that budding yeast Rif1 has unprecedented effects for a protein, inhibiting the recruitment of checkpoint proteins and RPA (Replication Protein A) to damaged chromosome regions, without significantly affecting the accumulation of ssDNA at those regions. Using chromatin immuno-precipitation, we provide evidence that Rif1 acts as a molecular "band-aid" for ssDNA lesions, associating with DNA damage independently of Rap1. In consequence, small or incipient lesions are protected from RPA and checkpoint proteins. When longer stretches of ssDNA are generated, they extend beyond the junction-proximal Rif1-protected regions. In consequence, the damage is detected and checkpoint signals are fired, resulting in cell cycle arrest. However, increased Rif1 expression raises the checkpoint-activation threshold to the point it simulates a checkpoint knockout and can also terminate a checkpoint arrest, despite persistent telomere deficiency. Our work has important implications for understanding the checkpoint and RPA-dependent DNA-damage responses in eukaryotic cells.
authors.affiliation: Institute for Ageing and Health, Newcastle University, Newcastle upon Tyne, United Kingdom.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Xue
authors.forename: Michael D
authors.initials: MD
authors.lastname: Rushton
authors.forename: Laura
authors.initials: L
authors.lastname: Maringele
mesh_terms: D059447:Cell Cycle Checkpoints / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002875:Chromosomes / Q000235:genetics; D004249:DNA Damage / Q000235:genetics; D004260:DNA Repair / Q000235:genetics; D004261:DNA Replication / Q000235:genetics; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D051716:Replication Protein A / Q000235:genetics / Q000378:metabolism*; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D000089804:Shelterin Complex; D016615:Telomere / Q000235:genetics; D034501:Telomere-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D004277:DNA, Single-Stranded; C440829:RAP1 protein, S cerevisiae; C082728:RFA1 protein, S cerevisiae; D051716:Replication Protein A; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D000089804:Shelterin Complex; D034501:Telomere-Binding Proteins; D014157:Transcription Factors; C074461:RIF1 protein, S cerevisiae