Early initiation of a replication origin tethered at the nuclear periphery.
Peripheral nuclear localization of chromosomal loci correlates with late replication in yeast and metazoan cells. To test whether peripheral positioning can impose late replication, we examined whether artificial tethering of an early-initiating replication origin to the nuclear periphery delays its replication in budding yeast. We tested the effects of three different peripheral tethering constructs on the time of replication of the early replication origin ARS607. Using the dense-isotope transfer method to assess replication time, we found that ARS607 still replicates early when tethered to the nuclear periphery using the Yif1 protein or a fragment of Sir4, whereas tethering using a Yku80 construct produces only a very slight replication delay. Single-cell microscopic analysis revealed no correlation between peripheral positioning of ARS607 in individual cells and delayed replication. Overall, our results demonstrate that a replication origin can initiate replication early in S phase, even if artificially relocated to the nuclear periphery.
Ebrahimi
Hani
H
Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD, UK.
Robertson
E Douglas
ED
Taddei
Angela
A
Gasser
Susan M
SM
Donaldson
Anne D
AD
Hiraga
Shin-ichiro
S
D033942:Adaptor Proteins, Vesicular Transport; D002467:Cell Nucleus / Q000235:genetics* / Q000378:metabolism; D042002:Chromatin Assembly and Disassembly / Q000235:genetics; D004261:DNA Replication; D004271:DNA, Fungal / Q000032:analysis; D004268:DNA-Binding Proteins / Q000235:genetics; D006056:Golgi Apparatus / Q000378:metabolism*; D008853:Microscopy; D015202:Protein Engineering; D021382:Protein Sorting Signals / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D018741:Replication Origin / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D004718:Saccharomycetales / Q000235:genetics*; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae / Q000235:genetics
D033942:Adaptor Proteins, Vesicular Transport; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; C061573:SIR4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; C473803:YIF1 protein, S cerevisiae; C520770:YKU80 protein, S cerevisiae