scholarly journals Chromatin loading of MCM hexamers is associated with di-/tri-methylation of histone H4K20 toward S phase entry

2021 ◽  
Author(s):  
Yoko Hayashi-Takanaka ◽  
Yuichiro Hayashi ◽  
Yasuhiro Hirano ◽  
Atsuko Miyawaki-Kuwakado ◽  
Yasuyuki Ohkawa ◽  
...  
Keyword(s):  
Dna Replication ◽  
S Phase ◽  
G1 Phase ◽  
Histone H4 ◽  
Replication Origins ◽  
Minichromosome Maintenance ◽  
Limiting Step ◽  
Mcm Helicase ◽  
Dna Replication Origins ◽  
Phase Entry

Replication of genomic DNA is a key step in initiating cell proliferation. Loading hexameric complexes of minichromosome maintenance (MCM) helicase on DNA replication origins during the G1 phase is essential in initiating DNA replication. Here, we show that stepwise loading of two hexamer complexes of MCM occurs during G1 progression in human cells. This transition from the single-to-double hexamer was associated with levels of methylation at lysine 20 of histone H4 (H4K20). A single hexamer of MCM complexes was loaded at the replication origins with the presence of H4K20 monomethylation (H4K20me1) in the early G1 phase, then another single hexamer was recruited to form a double hexamer later in G1 as H4K20me1 was converted to di-/tri-methylation (H4K20me2/me3). Under non-proliferating conditions, cells stay halted at the single-hexamer state in the presence of H4K20me1. We propose that the single-hexamer state on chromatin is a limiting step in making the proliferation-quiescence decision.

scholarly journals Cdc6 ATPase activity disengages Cdc6 from the pre-replicative complex to promote DNA replication

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
FuJung Chang ◽  
Alberto Riera ◽  
Cecile Evrin ◽  
Jingchuan Sun ◽  
Huilin Li ◽  
...  
Keyword(s):  
Dna Replication ◽  
Atpase Activity ◽  
Atp Hydrolysis ◽  
S Phase ◽  
G1 Phase ◽  
Helicase Loading ◽  
Mcm Helicase

To initiate DNA replication, cells first load an MCM helicase double hexamer at origins in a reaction requiring ORC, Cdc6, and Cdt1, also called pre-replicative complex (pre-RC) assembly. The essential mechanistic role of Cdc6 ATP hydrolysis in this reaction is still incompletely understood. Here, we show that although Cdc6 ATP hydrolysis is essential to initiate DNA replication, it is not essential for MCM loading. Using purified proteins, an ATPase-defective Cdc6 mutant ‘Cdc6-E224Q’ promoted MCM loading on DNA. Cdc6-E224Q also promoted MCM binding at origins in vivo but cells remained blocked in G1-phase. If after loading MCM, Cdc6-E224Q was degraded, cells entered an apparently normal S-phase and replicated DNA, a phenotype seen with two additional Cdc6 ATPase-defective mutants. Cdc6 ATP hydrolysis is therefore required for Cdc6 disengagement from the pre-RC after helicase loading to advance subsequent steps in helicase activation in vivo.

scholarly journals Intrinsic checkpoint deficiency during cell cycle re-entry from quiescence

2019 ◽  
Vol 218 (7) ◽  
pp. 2169-2184 ◽  
Author(s):  
Jacob Peter Matson ◽  
Amy M. House ◽  
Gavin D. Grant ◽  
Huaitong Wu ◽  
Joanna Perez ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Live Cell Imaging ◽  
Genome Instability ◽  
Replication Stress ◽  
S Phase ◽  
Minichromosome Maintenance ◽  
Cell Cycle Entry ◽  
Origin Licensing ◽  
Dna Replication Origins ◽  
Phase Entry

To maintain tissue homeostasis, cells transition between cell cycle quiescence and proliferation. An essential G1 process is minichromosome maintenance complex (MCM) loading at DNA replication origins to prepare for S phase, known as origin licensing. A p53-dependent origin licensing checkpoint normally ensures sufficient MCM loading before S phase entry. We used quantitative flow cytometry and live cell imaging to compare MCM loading during the long first G1 upon cell cycle entry and the shorter G1 phases in the second and subsequent cycles. We discovered that despite the longer G1 phase, the first G1 after cell cycle re-entry is significantly underlicensed. Consequently, the first S phase cells are hypersensitive to replication stress. This underlicensing results from a combination of slow MCM loading with a severely compromised origin licensing checkpoint. The hypersensitivity to replication stress increases over repeated rounds of quiescence. Thus, underlicensing after cell cycle re-entry from quiescence distinguishes a higher-risk first cell cycle that likely promotes genome instability.

scholarly journals Mechanism of Archaeal MCM Helicase Recruitment to DNA Replication Origins

2016 ◽  
Vol 61 (2) ◽  
pp. 287-296 ◽  
Author(s):  
Rachel Y. Samson ◽  
Priyanka D. Abeyrathne ◽  
Stephen D. Bell
Keyword(s):  
Dna Replication ◽  
Replication Origins ◽  
Mcm Helicase ◽  
Dna Replication Origins

scholarly journals Cohesin-mediated loop anchors confine the location of human replication origins

2021 ◽  
Author(s):  
Daniel Emerson ◽  
Peiyao A Zhao ◽  
Kyle Klein ◽  
Chunmin Ge ◽  
Linda Zhou ◽  
...  
Keyword(s):  
Dna Replication ◽  
Binding Sites ◽  
Genome Stability ◽  
S Phase ◽  
Replication Initiation ◽  
G1 Phase ◽  
Replication Origins ◽  
Topologically Associating Domains ◽  
Molecular Events ◽  
Dividing Cells

AbstractDNA replication occurs through an intricately regulated series of molecular events and is fundamental for genome stability across dividing cells in metazoans. It is currently unknown how the location of replication origins and the timing of their activation is determined in the human genome. Here, we dissect the role for G1 phase topologically associating domains (TADs), subTADs, and loops in the activation of replication initiation zones (IZs). We identify twelve subtypes of self-interacting chromatin domains distinguished by their degree of nesting, the presence of corner dot structures indicative of loops, and their co-localization with A/B compartments. Early replicating IZs localize to boundaries of nested corner-dot TAD/subTADs anchored by high density arrays of co-occupied CTCF+cohesin binding sites with divergently oriented motifs. By contrast, late replicating IZs localize to weak TADs/subTAD boundaries devoid of corner dots and most often anchored by singlet CTCF+cohesin sites. Upon global knock-down of cohesin-mediated loops in G1, early wave focal IZs replicate later in S phase and convert to diffuse placement along the genome. Moreover, IZs in mid-late S phase are delayed to the final minutes before entry into G2 when cohesin-mediated dot-less boundaries are ablated. We also delete a specific loop anchor and observe a sharp local delay of an early wave IZ to replication in late S phase. Our data demonstrate that cohesin-mediated loops at genetically-encoded TAD/subTAD boundaries in G1 phase are an essential determinant of the precise genomic placement of human replication origins in S phase.

scholarly journals Effect of minichromosome maintenance protein 2 deficiency on the locations of DNA replication origins

2015 ◽  
Vol 25 (4) ◽  
pp. 558-569 ◽  
Author(s):  
Dimiter Kunnev ◽  
Amy Freeland ◽  
Maochun Qin ◽  
Robert W. Leach ◽  
Jianmin Wang ◽  
...  
Keyword(s):  
Dna Replication ◽  
Replication Origins ◽  
Minichromosome Maintenance ◽  
Minichromosome Maintenance Protein ◽  
Dna Replication Origins

scholarly journals Rapid DNA replication origin licensing protects stem cell pluripotency

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Jacob Peter Matson ◽  
Raluca Dumitru ◽  
Philip Coryell ◽  
Ryan M Baxley ◽  
Weili Chen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Dna Replication ◽  
Cell Types ◽  
Minichromosome Maintenance ◽  
Origin Licensing ◽  
Mcm Helicase ◽  
Dna Replication Origin ◽  
Stem Cell Pluripotency ◽  
Dna Replication Origins ◽  
Intrinsic Characteristic

Complete and robust human genome duplication requires loading minichromosome maintenance (MCM) helicase complexes at many DNA replication origins, an essential process termed origin licensing. Licensing is restricted to G1 phase of the cell cycle, but G1 length varies widely among cell types. Using quantitative single-cell analyses, we found that pluripotent stem cells with naturally short G1 phases load MCM much faster than their isogenic differentiated counterparts with long G1 phases. During the earliest stages of differentiation toward all lineages, MCM loading slows concurrently with G1 lengthening, revealing developmental control of MCM loading. In contrast, ectopic Cyclin E overproduction uncouples short G1 from fast MCM loading. Rapid licensing in stem cells is caused by accumulation of the MCM loading protein, Cdt1. Prematurely slowing MCM loading in pluripotent cells not only lengthens G1 but also accelerates differentiation. Thus, rapid origin licensing is an intrinsic characteristic of stem cells that contributes to pluripotency maintenance.

Efficiency and equity in origin licensing to ensure complete DNA replication

2021 ◽  
Author(s):  
Liu Mei ◽  
Jeanette Gowen Cook
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Dna Replication ◽  
Genome Stability ◽  
S Phase ◽  
Replication Initiation ◽  
G1 Phase ◽  
Dna Replication Initiation ◽  
Origin Licensing ◽  
Dna Replication Origins

The cell division cycle must be strictly regulated during both development and adult maintenance, and efficient and well-controlled DNA replication is a key event in the cell cycle. DNA replication origins are prepared in G1 phase of the cell cycle in a process known as origin licensing which is essential for DNA replication initiation in the subsequent S phase. Appropriate origin licensing includes: (1) Licensing enough origins at adequate origin licensing speed to complete licensing before G1 phase ends; (2) Licensing origins such that they are well-distributed on all chromosomes. Both aspects of licensing are critical for replication efficiency and accuracy. In this minireview, we will discuss recent advances in defining how origin licensing speed and distribution are critical to ensure DNA replication completion and genome stability.

scholarly journals Identification of two telomere-proximal fission yeast DNA replication origins constrained by nearby cis-acting sequences to replicate in late S phase

F1000Research ◽  
2012 ◽  
Vol 1 ◽  
pp. 58 ◽  
Author(s):  
Amna Chaudari ◽  
Joel A Huberman
Keyword(s):  
Dna Replication ◽  
Fission Yeast ◽  
S Phase ◽  
Yeast Genome ◽  
Late Replication ◽  
Replication Origins ◽  
Replication Checkpoint ◽  
Cis Acting ◽  
Associated Sequences ◽  
Dna Replication Origins

Telomeres of the fission yeast, Schizosaccharomyces pombe, are known to replicate in late S phase, but the reasons for this late replication are not fully understood. We have identified two closely-spaced DNA replication origins, 5.5 to 8 kb upstream from the telomere itself. These are the most telomere-proximal of all the replication origins in the fission yeast genome. When located by themselves in circular plasmids, these origins fired in early S phase, but if flanking sequences closer to the telomere were included in the circular plasmid, then replication was restrained to late S phase – except in cells lacking the replication-checkpoint kinase, Cds1. We conclude that checkpoint-dependent late replication of telomere-associated sequences is dependent on nearby cis-acting sequences, not on proximity to the physical end of a linear chromosome.

scholarly journals Intrinsic checkpoint deficiency during cell cycle re-entry from quiescence

2019 ◽  
Author(s):  
Jacob Peter Matson ◽  
Amy M. House ◽  
Gavin D. Grant ◽  
Huaitong Wu ◽  
Joanna Perez ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Genome Instability ◽  
Replication Stress ◽  
S Phase ◽  
Minichromosome Maintenance ◽  
Cell Cycle Entry ◽  
Origin Licensing ◽  
Dna Replication Origin ◽  
Dna Replication Origins

SUMMARYThe authors find that human cells re-entering the cell cycle from quiescence have both an impaired p53-dependent DNA replication origin licensing checkpoint and slow origin licensing. This combination makes every first S phase underlicensed and hypersensitive to replication stress.ABSTRACTTo maintain tissue homeostasis, cells transition between cell cycle quiescence and proliferation. An essential G1 process is Minichromosome Maintenance complex (MCM) loading at DNA replication origins to prepare for S phase, known as origin licensing. A p53-dependent origin licensing checkpoint normally ensures sufficient MCM loading prior to S phase entry. We used quantitative flow cytometry and live cell imaging to compare MCM loading during the long first G1 upon cell cycle entry and the shorter G1 phases in the second and subsequent cycles. We discovered that despite the longer G1 phase, the first G1 after cell cycle re-entry is significantly underlicensed. As a result, the first S phase cells are hypersensitive to replication stress. This underlicensing is from a combination of slow MCM loading with a severely compromised origin licensing checkpoint. The hypersensitivity to replication stress increases over repeated rounds of quiescence. Thus, underlicensing after cell cycle re-entry from quiescence distinguishes a higher risk cell cycle that promotes genome instability.

scholarly journals Yra1 Is Required for S Phase Entry and Affects Dia2 Binding to Replication Origins

2007 ◽  
Vol 27 (13) ◽  
pp. 4674-4684 ◽  
Author(s):  
Swarna Swaminathan ◽  
Andrew C. Kile ◽  
Elizabeth M. MacDonald ◽  
Deanna M. Koepp
Keyword(s):  
Dna Replication ◽  
Mrna Export ◽  
S Phase ◽  
Temperature Sensitive ◽  
Replication Origins ◽  
Expression Levels ◽  
C Terminus ◽  
Rna Export ◽  
F Box Protein ◽  
Phase Entry

ABSTRACT The Saccharomyces cerevisiae F-box protein Dia2 is important for DNA replication and genomic stability. Using an affinity approach, we identified Yra1, a transcription-coupled mRNA export protein, as a Dia2 interaction partner. We find that yra1 mutants are sensitive to DIA2 expression levels. Like Dia2, Yra1 associates with chromatin and binds replication origins, suggesting that they may function together in DNA replication. Consistent with this idea, Yra1 and Dia2 coimmunoprecipitate with Hys2, a subunit of DNA polymerase δ. The C terminus of Yra1 is required to interact with Dia2. A yra1 mutant that lacks this domain is temperature sensitive yet has no apparent defect in RNA export. Remarkably, this mutant also fails to enter S phase at the nonpermissive temperature. Significantly, other mutants in transcription-coupled export do not exhibit S phase entry defects or sensitivity to DIA2 expression levels. Together, these results indicate that Yra1 has a role in DNA replication distinct from its role in mRNA export. Furthermore, Dia2 binding to replication origins is significantly reduced when association with Yra1 is compromised, suggesting that one aspect of the role of Yra1 in DNA replication may involve recruiting Dia2 to chromatin.

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