scholarly journals The effects of manipulating levels of replication initiation factors on origin firing efficiency in yeast

PLoS Genetics ◽  
2019 ◽  
Vol 15 (10) ◽  
pp. e1008430 ◽  
Author(s):  
Kelsey L. Lynch ◽  
Gina M. Alvino ◽  
Elizabeth X. Kwan ◽  
Bonita J. Brewer ◽  
M. K. Raghuraman
Keyword(s):  
Replication Initiation ◽  
Origin Firing ◽  
Initiation Factors ◽  
Firing Efficiency

scholarly journals Checkpoint inhibition of origin firing prevents inappropriate replication outside of S-phase

2020 ◽  
Author(s):  
Mark C. Johnson ◽  
Geylani Can ◽  
Miguel Santos ◽  
Diana Alexander ◽  
Philip Zegerman
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
S Phase ◽  
Dna Lesions ◽  
Replication Initiation ◽  
Origin Firing ◽  
Initiation Factors ◽  
Dependent Inhibition ◽  
Rad53 Kinase ◽  
S Phase Checkpoint

AbstractAcross eukaryotes, checkpoints maintain the order of cell cycle events in the face of DNA damage or incomplete replication. Although a wide array of DNA lesions activates the checkpoint kinases, whether and how this response differs in different phases of the cell cycle remains poorly understood. The S-phase checkpoint for example results in the slowing of replication, which in the budding yeast Saccharomyces cerevisiae is caused by Rad53 kinase-dependent inhibition of the initiation factors Sld3 and Dbf4. Despite this, we show here that Rad53 phosphorylates both of these substrates throughout the cell cycle at the same sites as in S-phase, suggesting roles for this pathway beyond S-phase. Indeed we show that Rad53-dependent inhibition of Sld3 and Dbf4 limits re-replication in G2/M phase, preventing inappropriate gene amplification events. In addition we show that inhibition of Sld3 and Dbf4 after DNA damage in G1 phase prevents premature replication initiation at all origins at the G1/S transition. This study redefines the scope and specificity of the ‘S-phase checkpoint’ with implications for understanding the roles of this checkpoint in the majority of cancers that lack proper cell cycle controls.

scholarly journals Limiting replication initiation factors execute the temporal programme of origin firing in budding yeast

2011 ◽  
Vol 30 (23) ◽  
pp. 4805-4814 ◽  
Author(s):  
Davide Mantiero ◽  
Amanda Mackenzie ◽  
Anne Donaldson ◽  
Philip Zegerman
Keyword(s):  
Budding Yeast ◽  
Replication Initiation ◽  
Origin Firing ◽  
Initiation Factors

scholarly journals The limiting DNA replication initiation factors Sld2 and Sld3 influence origin efficiency independent of origin firing time

2018 ◽  
Author(s):  
Kelsey L. Lynch ◽  
Elizabeth X. Kwan ◽  
Gina M. Alvino ◽  
Bonita J. Brewer ◽  
M.K. Raghuraman
Keyword(s):  
Rdna Locus ◽  
S Phase ◽  
Replication Initiation ◽  
Yeast Genome ◽  
Phase Duration ◽  
Origin Firing ◽  
Dna Replication Initiation ◽  
Initiation Factors ◽  
Origin Activation ◽  
Time Of Origin

AbstractChromosome replication in Saccharomyces cerevisiae is initiated from roughly 300 origins that are regulated both by DNA sequence and by the limited abundance of four trans-acting initiation proteins (Sld2, Sld3, Dpb11 and Dbf4, collectively called “SSDD”). We set out to determine how the association of Sld2 or Sld3 at origins contributes to time of origin activation and/or origin efficiency using auxin-induced protein degradation to further decrease their abundance. Depleting cells of either factor slows growth rate, increases S-phase duration, and causes viability defects, without activating the S phase checkpoint. Chr XII is uniquely unstable with breakage occurring specifically within the rDNA locus. The efficiency of the rDNA origin is decreased while the onset of replication initiation is unchanged. We found that origin efficiency is reduced uniformly across the unique portions of the yeast genome. We conclude that the abundance of Sld2 and Sld3 contribute primarily to origin efficiency.

scholarly journals Structural Changes in Mcm5 Protein Bypass Cdc7-Dbf4 Function and Reduce Replication Origin Efficiency in Saccharomyces cerevisiae

2007 ◽  
Vol 27 (21) ◽  
pp. 7594-7602 ◽  
Author(s):  
Margaret L. Hoang ◽  
Ronald P. Leon ◽  
Luis Pessoa-Brandao ◽  
Sonia Hunt ◽  
M. K. Raghuraman ◽  
...  
Keyword(s):  
Structural Changes ◽  
S Phase ◽  
Minichromosome Maintenance ◽  
Chromosomal Replication ◽  
Origin Firing ◽  
Highly Active ◽  
Mcm Complex ◽  
Alternate States ◽  
Firing Efficiency ◽  
Intrinsic Firing

ABSTRACT Eukaryotic chromosomal replication is a complicated process with many origins firing at different efficiencies and times during S phase. Prereplication complexes are assembled on all origins in G1 phase, and yet only a subset of complexes is activated during S phase by DDK (for Dbf4-dependent kinase) (Cdc7-Dbf4). The yeast mcm5-bob1 (P83L) mutation bypasses DDK but results in reduced intrinsic firing efficiency at 11 endogenous origins and at origins located on minichromosomes. Origin efficiency may result from Mcm5 protein assuming an altered conformation, as predicted from the atomic structure of an archaeal MCM (for minichromosome maintenance) homologue. Similarly, an intragenic mutation in a residue predicted to interact with P83L suppresses the mcm5-bob1 bypass phenotype. We propose DDK phosphorylation of the MCM complex normally results in a single, highly active conformation of Mcm5, whereas the mcm5-bob1 mutation produces a number of conformations, only one of which is permissive for origin activation. Random adoption of these alternate states by the mcm5-bob1 protein can explain both how origin firing occurs independently of DDK and why origin efficiency is reduced. Because similar mutations in mcm2 and mcm4 cannot bypass DDK, Mcm5 protein may be a unique Mcm protein that is the final target of DDK regulation.

scholarly journals Evolution of DNA Replication Origin Specification and Gene Silencing Mechanisms

2020 ◽  
Author(s):  
Y. Hu ◽  
A. Tareen ◽  
Y-J. Sheu ◽  
W. T. Ireland ◽  
C. Speck ◽  
...  
Keyword(s):  
Dna Replication ◽  
Gene Silencing ◽  
Origin Recognition Complex ◽  
Replication Initiation ◽  
Separate Analysis ◽  
Genome Replication ◽  
Sequence Specificity ◽  
Sequence Motifs ◽  
Origin Firing ◽  
Α Helix

AbstractDNA replication in eukaryotic cells initiates from chromosomal locations, called replication origins, that bind the Origin Recognition Complex (ORC) prior to S phase. Origin establishment is guided by well-defined DNA sequence motifs in Saccharomyces cerevisiae and some other budding yeasts, but most eukaryotes lack sequence-specific origins. At present, the mechanistic and evolutionary reasons for this difference are unclear. A 3.9 Å structure of S. cerevisiae ORC-Cdc6-Cdt1-Mcm2-7 (OCCM) bound to origin DNA revealed, among other things, that a loop within Orc2 inserts into a DNA minor groove and an α-helix within Orc4 inserts into a DNA major groove1. We show that this Orc4 α-helix mediates the sequence-specificity of origins in S. cerevisiae. Specifically, mutations were identified within this α-helix that alter the sequence-dependent activity of individual origins as well as change global genomic origin firing patterns. This was accomplished using a massively parallel origin selection assay analyzed using a custom mutual-information-based modeling approach and a separate analysis of whole-genome replication profiling and statistics. Interestingly, the sequence specificity of DNA replication initiation, as mediated by the Orc4 α-helix, has evolved in close conjunction with the gain of ORC-Sir4-mediated gene silencing and the loss of RNA interference.

scholarly journals DNA Replication Origins Fire Stochastically in Fission Yeast

2006 ◽  
Vol 17 (1) ◽  
pp. 308-316 ◽  
Author(s):  
Prasanta K. Patel ◽  
Benoit Arcangioli ◽  
Stephen P. Baker ◽  
Aaron Bensimon ◽  
Nicholas Rhind
Keyword(s):  
Dna Replication ◽  
Fission Yeast ◽  
Replication Initiation ◽  
Epigenetic Mechanism ◽  
Replication Origins ◽  
Origin Firing ◽  
Cell Cycles ◽  
Initiation Of Replication ◽  
Dna Combing ◽  
Dna Replication Origins

DNA replication initiates at discrete origins along eukaryotic chromosomes. However, in most organisms, origin firing is not efficient; a specific origin will fire in some but not all cell cycles. This observation raises the question of how individual origins are selected to fire and whether origin firing is globally coordinated to ensure an even distribution of replication initiation across the genome. We have addressed these questions by determining the location of firing origins on individual fission yeast DNA molecules using DNA combing. We show that the firing of replication origins is stochastic, leading to a random distribution of replication initiation. Furthermore, origin firing is independent between cell cycles; there is no epigenetic mechanism causing an origin that fires in one cell cycle to preferentially fire in the next. Thus, the fission yeast strategy for the initiation of replication is different from models of eukaryotic replication that propose coordinated origin firing.

Sign in / Sign up

Export Citation Format

Share Document