G2 phase chromatin lacks determinants of replication timing

DNA replication in all eukaryotes follows a defined replication timing program, the molecular mechanism of which remains elusive. Using a Xenopus laevis egg extract replication system, we previously demonstrated that replication timing is established during early G1 phase of the cell cycle (timing decision point [TDP]), which is coincident with the repositioning and anchorage of chromatin in the newly formed nucleus. In this study, we use this same system to show that G2 phase chromatin lacks determinants of replication timing but maintains the overall spatial organization of chromatin domains, and we confirm this finding by genome-wide analysis of rereplication in vivo. In contrast, chromatin from quiescent cells retains replication timing but exhibits disrupted spatial organization. These data support a model in which events at the TDP, facilitated by chromatin spatial organization, establish determinants of replication timing that persist independent of spatial organization until the process of chromatin replication during S phase erases those determinants.

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Faculty Opinions recommendation of Genome-wide analysis in vivo of translation with nucleotide resolution using ribosome profiling.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1148255.793503924 ◽

2015 ◽

Author(s):

Timothy T Weil

Keyword(s):

Ribosome Profiling ◽

Genome Wide Analysis ◽

Genome Wide ◽

Nucleotide Resolution

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Faculty Opinions recommendation of Genome-wide analysis in vivo of translation with nucleotide resolution using ribosome profiling.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1148255.626621 ◽

2009 ◽

Author(s):

Michael Kiebler ◽

Lucia Schoderböck

Keyword(s):

Ribosome Profiling ◽

Genome Wide Analysis ◽

Genome Wide ◽

Nucleotide Resolution

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Genome‐wide analysis of in vivo CcpA binding with and without its key co‐factor HPr in the major human pathogen group A Streptococcus

Molecular Microbiology ◽

10.1111/mmi.14667 ◽

2020 ◽

Author(s):

Sruti DebRoy ◽

Victor Aliaga‐Tobar ◽

Gabriel Galvez ◽

Srishtee Arora ◽

Xiaowen Liang ◽

...

Keyword(s):

Group A Streptococcus ◽

Human Pathogen ◽

Genome Wide Analysis ◽

Genome Wide ◽

Group A

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PGC7 suppresses TET3 for protecting DNA methylation

Nucleic Acids Research ◽

10.1093/nar/gkt1261 ◽

2013 ◽

Vol 42 (5) ◽

pp. 2893-2905 ◽

Cited By ~ 39

Author(s):

Chunjing Bian ◽

Xiaochun Yu

Keyword(s):

Dna Methylation ◽

Molecular Mechanism ◽

Biological Process ◽

Cpg Islands ◽

Binding Motifs ◽

Genome Wide Analysis ◽

Dna Motif ◽

Genome Wide

Abstract Ten-eleven translocation (TET) family enzymes convert 5-methylcytosine to 5-hydroxylmethylcytosine. However, the molecular mechanism that regulates this biological process is not clear. Here, we show the evidence that PGC7 (also known as Dppa3 or Stella) interacts with TET2 and TET3 both in vitro and in vivo to suppress the enzymatic activity of TET2 and TET3. Moreover, lacking PGC7 induces the loss of DNA methylation at imprinting loci. Genome-wide analysis of PGC7 reveals a consensus DNA motif that is recognized by PGC7. The CpG islands surrounding the PGC7-binding motifs are hypermethylated. Taken together, our study demonstrates a molecular mechanism by which PGC7 protects DNA methylation from TET family enzyme-dependent oxidation.

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Genome-Wide Analysis of the Arabidopsis Replication Timing Program

PLANT PHYSIOLOGY ◽

10.1104/pp.17.01537 ◽

2018 ◽

Vol 176 (3) ◽

pp. 2166-2185 ◽

Cited By ~ 13

Author(s):

Lorenzo Concia ◽

Ashley M. Brooks ◽

Emily Wheeler ◽

Gregory J. Zynda ◽

Emily E. Wear ◽

...

Keyword(s):

Replication Timing ◽

Genome Wide Analysis ◽

Genome Wide

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The viral Ki-ras gene must be expressed in the G2 phase if ts Kirsten sarcoma virus-infected NRK cells are to proliferate in serum-free medium

Molecular and Cellular Biology ◽

10.1128/mcb.7.1.444-449.1987 ◽

1987 ◽

Vol 7 (1) ◽

pp. 444-449

Author(s):

J P Durkin ◽

J F Whitfield

Keyword(s):

S Phase ◽

Temperature Sensitive ◽

Serum Free Medium ◽

Free Medium ◽

Ras Gene ◽

Serum Free ◽

Quiescent Cells ◽

Sarcoma Virus ◽

G2 Phase ◽

Oncogene Products

NRK cells infected with a temperature-sensitive Kirsten sarcoma virus (ts371 KSV) are transformed at 36 degrees C, but are untransformed at 41 degrees C which inactivates the abnormally thermolabile oncogenic p21Ki product of the viral Ki-ras gene. At 41 degrees C, tsKSV-infected NRK cells were arrested in G0/G1 when incubated in serum-free medium, but could then be stimulated to transit G1, replicate DNA, and divide by adding serum at 41 degrees C or dropping the temperature to a p21-activating 36 degrees C without adding serum. When quiescent cells at 41 degrees C were stimulated to transit G1 in serum-free medium by activating p21 at 36 degrees C and then shifted back to the p21-inactivating 41 degrees C in the mid-S phase, they continued replicating DNA but could not transit G2. Reactivating p21 in the G2-arrested cells by once again lowering the temperature to 36 degrees C stimulated a rapid entry into mitosis. By contrast, while serum-stimulated quiescent G0 cells at 41 degrees C replicate DNA and divide, serum did not induce G2-arrested cells to enter mitosis, indicating that serum growth factors may trigger events in the G1 phase that ultimately determine G2 transit. These observations made with the viral ras product suggest that cellular ras proto-oncogene products have a role in G2 transit of normal cells.

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Replication timing control can be maintained in extrachromosomally amplified genes.

Molecular and Cellular Biology ◽

10.1128/mcb.11.9.4779 ◽

1991 ◽

Vol 11 (9) ◽

pp. 4779-4785 ◽

Cited By ~ 13

Author(s):

S M Carroll ◽

J Trotter ◽

G M Wahl

Keyword(s):

Adenosine Deaminase ◽

Timing Analysis ◽

Single Gene ◽

Replication Timing ◽

Virus Genome ◽

S Phase ◽

Timing Control ◽

Barr Virus ◽

Extrachromosomal Elements

Extrachromosomal elements are common early intermediates of gene amplification in vivo and in cell culture. The time at which several extrachromosomal elements replicate was compared with that of the corresponding amplified or unamplified chromosomal sequences. The replication timing analysis employed a retroactive synchrony method in which fluorescence-activated cell sorting was used to obtain cells at different stages of the cell cycle. Extrachromosomally amplified Syrian hamster CAD genes (CAD is an acronym for the single gene which encodes the trifunctional protein which catalyzes the first three steps of uridine biosynthesis) replicated in a narrow window of early S-phase which was approximately the same as that of chromosomally amplified CAD genes. Similarly, extrachromosomally amplified mouse adenosine deaminase genes replicated at a discrete time in early S-phase which approximated the replication time of the unamplified adenosine deaminase gene. In contrast, the multicopy extrachromosomal Epstein-Barr virus genome replicated within a narrow window in late S-phase in latently infected human Rajii cells. The data indicate that localization within a chromosome is not required for the maintenance of replication timing control.

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Transcription-mediated organization of the replication initiation program across large genes sets up common fragile sites genome-wide

10.1101/714717 ◽

2019 ◽

Cited By ~ 1

Author(s):

Olivier Brison ◽

Sami EL-Hilali ◽

Dana Azar ◽

Stéphane Koundrioukoff ◽

Mélanie Schmidt ◽

...

Keyword(s):

Replication Timing ◽

S Phase ◽

Fragile Sites ◽

Replication Initiation ◽

Common Fragile Sites ◽

Large Gene ◽

Genome Wide ◽

Underlying Mechanisms ◽

Large Genes ◽

Nascent Transcripts

ABSTRACTCommon Fragile Sites (CFSs) are chromosome regions prone to breakage under replication stress, known to drive chromosome rearrangements during oncogenesis. Most CFSs nest in large expressed genes, suggesting that transcription elicits their instability but the underlying mechanisms remained elusive. Analyses of genome-wide replication timing of human lymphoblasts here show that stress-induced delayed/under-replication is the hallmark of CFSs. Extensive genome-wide analyses of nascent transcripts, replication origin positioning and fork directionality reveal that 80% of CFSs nest in large transcribed domains poor in initiation events, thus replicated by long-traveling forks. In contrast to formation of sequence-dependent fork barriers or head-on transcription-replication conflicts, traveling-long in late S phase explains CFS replication features. We further show that transcription inhibition during the S phase, which excludes the setting of new replication origins, fails to rescue CFS stability. Altogether, results show that transcription-dependent suppression of initiation events delays replication of large gene body, committing them to instability.

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