scholarly journals Transcription-coupled structural dynamics of topologically associating domains regulate replication origin efficiency

2020 ◽  
Author(s):  
Yongzheng Li ◽  
Boxin Xue ◽  
Liwei Zhang ◽  
Qian Peter Su ◽  
Mengling Zhang ◽  
...  
Keyword(s):  
Dna Replication ◽  
Structural Dynamics ◽  
Super Resolution ◽  
S Phase ◽  
Replication Origins ◽  
Topologically Associating Domains ◽  
Chromatin Structural ◽  
Resolution Imaging ◽  
Epigenetic Signatures ◽  
Selection Of

ABSTRACTMetazoan cells only utilize a small subset of the potential DNA replication origins to duplicate the whole genome in each cell cycle. Origin choice is linked to cell growth, differentiation, and replication stress. Despite various genetic and epigenetic signatures are found to be related with active origins, it remains elusive how the selection of origins is determined. The classic Rosette model proposes that the origins clustered in a chromatin domain are preferentially and simultaneously fired, but direct imaging evidence has been lacking due to insufficient spatial resolution. Here, we applied dual-color stochastic optical reconstruction microscopy (STORM) super-resolution imaging to map the spatial distribution of origins within individual topologically associating domains (TADs). We found that multiple replication origins initiate separately at the spatial boundary of a TAD at the beginning of the S phase, in contrary to the Rosette model. Intriguingly, while both active and dormant origins are distributed homogeneously in the TAD during the G1 phase, active origins relocate to the TAD periphery before entering the S phase. We proved that such origin relocalization is dependent on both transcription and CTCF-mediated chromatin structure. Further, we observed that the replication machinery protein PCNA forms immobile clusters around the TADs at the G1/S transition, which explains why origins at the TAD periphery are preferentially fired. Thus, we propose a “Chromatin Re-organization Induced Selective Initiation” (CRISI) model that the transcription-coupled chromatin structural re-organization determines the selection of replication origins, which transcends the scope of specific genetic and epigenetic signatures for origin efficiency. Our in situ super-resolution imaging unveiled coordination among DNA replication, transcription, and chromatin organization inside individual TADs, providing new insights into the biological functions of sub-domain chromatin structural dynamics.

scholarly journals Transcription-coupled structural dynamics of topologically associating domains regulate replication origin efficiency

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Yongzheng Li ◽  
Boxin Xue ◽  
Mengling Zhang ◽  
Liwei Zhang ◽  
Yingping Hou ◽  
...  
Keyword(s):  
Dna Replication ◽  
Structural Dynamics ◽  
Replication Origin ◽  
High Efficiency ◽  
S Phase ◽  
Chromatin Domain ◽  
Replication Origins ◽  
Replication Efficiency ◽  
Topologically Associating Domains ◽  
Epigenetic Signatures

Abstract Background Metazoan cells only utilize a small subset of the potential DNA replication origins to duplicate the whole genome in each cell cycle. Origin choice is linked to cell growth, differentiation, and replication stress. Although various genetic and epigenetic signatures have been linked to the replication efficiency of origins, there is no consensus on how the selection of origins is determined. Results We apply dual-color stochastic optical reconstruction microscopy (STORM) super-resolution imaging to map the spatial distribution of origins within individual topologically associating domains (TADs). We find that multiple replication origins initiate separately at the spatial boundary of a TAD at the beginning of the S phase. Intriguingly, while both high-efficiency and low-efficiency origins are distributed homogeneously in the TAD during the G1 phase, high-efficiency origins relocate to the TAD periphery before the S phase. Origin relocalization is dependent on both transcription and CTCF-mediated chromatin structure. Further, we observe that the replication machinery protein PCNA forms immobile clusters around TADs at the G1/S transition, explaining why origins at the TAD periphery are preferentially fired. Conclusion Our work reveals a new origin selection mechanism that the replication efficiency of origins is determined by their physical distribution in the chromatin domain, which undergoes a transcription-dependent structural re-organization process. Our model explains the complex links between replication origin efficiency and many genetic and epigenetic signatures that mark active transcription. The coordination between DNA replication, transcription, and chromatin organization inside individual TADs also provides new insights into the biological functions of sub-domain chromatin structural dynamics.

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 Superresolution imaging reveals spatiotemporal propagation of human replication foci mediated by CTCF-organized chromatin structures

2020 ◽  
Vol 117 (26) ◽  
pp. 15036-15046 ◽  
Author(s):  
Qian Peter Su ◽  
Ziqing Winston Zhao ◽  
Luming Meng ◽  
Miao Ding ◽  
Weiwei Zhang ◽  
...  
Keyword(s):  
Dna Replication ◽  
S Phase ◽  
Replication Origins ◽  
Nuclear Dynamics ◽  
Superresolution Imaging ◽  
Stochastic Optical Reconstruction Microscopy ◽  
Replication Foci ◽  
Optical Reconstruction ◽  
Epigenetic Signatures

Mammalian DNA replication is initiated at numerous replication origins, which are clustered into thousands of replication domains (RDs) across the genome. However, it remains unclear whether the replication origins within each RD are activated stochastically or preferentially near certain chromatin features. To understand how DNA replication in single human cells is regulated at the sub-RD level, we directly visualized and quantitatively characterized the spatiotemporal organization, morphology, and in situ epigenetic signatures of individual replication foci (RFi) across S-phase at superresolution using stochastic optical reconstruction microscopy. Importantly, we revealed a hierarchical radial pattern of RFi propagation dynamics that reverses directionality from early to late S-phase and is diminished upon caffeine treatment or CTCF knockdown. Together with simulation and bioinformatic analyses, our findings point to a “CTCF-organized REplication Propagation” (CoREP) model, which suggests a nonrandom selection mechanism for replication activation at the sub-RD level during early S-phase, mediated by CTCF-organized chromatin structures. Collectively, these findings offer critical insights into the key involvement of local epigenetic environment in coordinating DNA replication across the genome and have broad implications for our conceptualization of the role of multiscale chromatin architecture in regulating diverse cell nuclear dynamics in space and time.

scholarly journals CTCF-mediated Chromatin Structures Dictate the Spatio-temporal Propagation of Replication Foci

2019 ◽  
Author(s):  
Qian Peter Su ◽  
Ziqing Winston Zhao ◽  
Luming Meng ◽  
Miao Ding ◽  
Weiwei Zhang ◽  
...  
Keyword(s):  
Super Resolution ◽  
S Phase ◽  
Temporal Organization ◽  
Replication Origins ◽  
Stochastic Optical Reconstruction Microscopy ◽  
Replication Foci ◽  
Spatio Temporal ◽  
Optical Reconstruction ◽  
Epigenetic Signatures

ABSTRACTMammalian DNA replication is initiated at numerous replication origins, which are clustered into thousands of replication domains (RDs) across the genome. However, it remains unclear whether the replication origins within each RD are activated stochastically. To understand how replication is regulated at the sub-RD level, we directly visualized the spatio-temporal organization, morphology, and in situ epigenetic signatures of individual replication foci (RFi) across S-phase using super-resolution stochastic optical reconstruction microscopy (STORM). Importantly, we revealed a hierarchical radial pattern of RFi propagation that reverses its directionality from early to late S-phase, and is diminished upon caffeine treatment or CTCF knockdown. Together with simulation and bioinformatic analyses, our findings point to a ‘CTCF-organized REplication Propagation’ (CoREP) model. The CoREP model suggests a non-random selection mechanism for replication activation mediated by CTCF at the sub-RD level, as well as the critical involvement of local chromatin environment in regulating replication in space and time.

scholarly journals Snapshots of archaeal DNA replication and repair in living cells using super-resolution imaging

2018 ◽  
Author(s):  
Floriane Delpech ◽  
Yoann Collien ◽  
Pierre Mahou ◽  
Emmanuel Beaurepaire ◽  
Hannu Myllykallio ◽  
...  
Keyword(s):  
Dna Replication ◽  
Super Resolution ◽  
Living Cells ◽  
Dna Replication And Repair ◽  
Resolution Imaging ◽  
Archaeal Dna Replication

scholarly journals The conserved bromo-adjacent homology domain of yeast Orc1 functions in the selection of DNA replication origins within chromatin

2010 ◽  
Vol 24 (13) ◽  
pp. 1418-1433 ◽  
Author(s):  
P. Muller ◽  
S. Park ◽  
E. Shor ◽  
D. J. Huebert ◽  
C. L. Warren ◽  
...  
Keyword(s):  
Dna Replication ◽  
Replication Origins ◽  
Dna Replication Origins ◽  
Selection Of

Chromosomal DNA replication initiates at the same origins in meiosis and mitosis

1994 ◽  
Vol 14 (5) ◽  
pp. 3524-3534
Author(s):  
I Collins ◽  
C S Newlon
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
S Phase ◽  
Replication Origins ◽  
Chromosomal Dna ◽  
Chromosomal Replication ◽  
Yeast Saccharomyces Cerevisiae ◽  
Autonomously Replicating Sequence ◽  
Ars Elements ◽  
Chromosome Iii

Autonomously replicating sequence (ARS) elements are identified by their ability to promote high-frequency transformation and extrachromosomal replication of plasmids in the yeast Saccharomyces cerevisiae. Six of the 14 ARS elements present in a 200-kb region of Saccharomyces cerevisiae chromosome III are mitotic chromosomal replication origins. The unexpected observation that eight ARS elements do not function at detectable levels as chromosomal replication origins during mitotic growth suggested that these ARS elements may function as chromosomal origins during premeiotic S phase. Two-dimensional agarose gel electrophoresis was used to map premeiotic replication origins in a 100-kb segment of chromosome III between HML and CEN3. The pattern of origin usage in premeiotic S phase was identical to that in mitotic S phase, with the possible exception of ARS308, which is an inefficient mitotic origin associated with CEN3. CEN3 was found to replicate during premeiotic S phase, demonstrating that the failure of sister chromatids to disjoin during the meiosis I division is not due to unreplicated centromeres. No origins were found in the DNA fragments without ARS function. Thus, in both mitosis and meiosis, chromosomal replication origins are coincident with ARS elements but not all ARS elements have chromosomal origin function. The efficiency of origin use and the patterns of replication termination are similar in meiosis and in mitosis. DNA replication termination occurs over a broad distance between active origins.

scholarly journals Chromosomal DNA replication initiates at the same origins in meiosis and mitosis.

1994 ◽  
Vol 14 (5) ◽  
pp. 3524-3534 ◽  
Author(s):  
I Collins ◽  
C S Newlon
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
S Phase ◽  
Replication Origins ◽  
Chromosomal Dna ◽  
Chromosomal Replication ◽  
Yeast Saccharomyces Cerevisiae ◽  
Autonomously Replicating Sequence ◽  
Ars Elements ◽  
Chromosome Iii

Autonomously replicating sequence (ARS) elements are identified by their ability to promote high-frequency transformation and extrachromosomal replication of plasmids in the yeast Saccharomyces cerevisiae. Six of the 14 ARS elements present in a 200-kb region of Saccharomyces cerevisiae chromosome III are mitotic chromosomal replication origins. The unexpected observation that eight ARS elements do not function at detectable levels as chromosomal replication origins during mitotic growth suggested that these ARS elements may function as chromosomal origins during premeiotic S phase. Two-dimensional agarose gel electrophoresis was used to map premeiotic replication origins in a 100-kb segment of chromosome III between HML and CEN3. The pattern of origin usage in premeiotic S phase was identical to that in mitotic S phase, with the possible exception of ARS308, which is an inefficient mitotic origin associated with CEN3. CEN3 was found to replicate during premeiotic S phase, demonstrating that the failure of sister chromatids to disjoin during the meiosis I division is not due to unreplicated centromeres. No origins were found in the DNA fragments without ARS function. Thus, in both mitosis and meiosis, chromosomal replication origins are coincident with ARS elements but not all ARS elements have chromosomal origin function. The efficiency of origin use and the patterns of replication termination are similar in meiosis and in mitosis. DNA replication termination occurs over a broad distance between active origins.

scholarly journals Changes in association of the Xenopus origin recognition complex with chromatin on licensing of replication origins

1999 ◽  
Vol 112 (12) ◽  
pp. 2011-2018 ◽  
Author(s):  
A. Rowles ◽  
S. Tada ◽  
J.J. Blow
Keyword(s):  
Dna Replication ◽  
Single Molecule ◽  
Replication Origin ◽  
Origin Recognition Complex ◽  
S Phase ◽  
Replication Origins ◽  
Free System ◽  
Essential Function ◽  
Initiation Of Dna Replication ◽  
Origin Recognition

During late mitosis and early G1, a series of proteins are assembled onto replication origins that results in them becoming ‘licensed’ for replication in the subsequent S phase. In Xenopus this first involves the assembly onto chromatin of the Xenopus origin recognition complex XORC, and then XCdc6, and finally the RLF-M component of the replication licensing system. In this paper we examine changes in the way that XORC associates with chromatin in the Xenopus cell-free system as origins become licensed. Restricting the quantity of XORC on chromatin reduced the extent of replication as expected if a single molecule of XORC is sufficient to specify a single replication origin. During metaphase, XOrc1 associated only weakly with chromatin. In early interphase, XOrc1 formed a strong complex with chromatin, as evidenced by its resistance to elution by 200 mM salt, and this state persisted when XCdc6 was assembled onto the chromatin. As a consequence of origins becoming licensed the association of XOrc1 and XCdc6 with chromatin was destabilised, and XOrc1 became susceptible to removal from chromatin by exposure to either high salt or high Cdk levels. At this stage the essential function for XORC and XCdc6 in DNA replication had already been fulfilled. Since high Cdk levels are required for the initiation of DNA replication, this ‘licensing-dependent origin inactivation’ may contribute to mechanisms that prevent re-licensing of replication origins once S phase has started.

scholarly journals The Human Stress-Activated Protein kin17 Belongs to the Multiprotein DNA Replication Complex and Associates In Vivo with Mammalian Replication Origins

2005 ◽  
Vol 25 (9) ◽  
pp. 3814-3830 ◽  
Author(s):  
Laurent Miccoli ◽  
Isabelle Frouin ◽  
Olivia Novac ◽  
Domenic Di Paola ◽  
Francis Harper ◽  
...  
Keyword(s):  
Dna Replication ◽  
S Phase ◽  
Dna Fragments ◽  
Replication Origins ◽  
Proliferating Cells ◽  
Direct Role ◽  
Replication Complex ◽  
Human Stress

ABSTRACT The human stress-activated protein kin17 accumulates in the nuclei of proliferating cells with predominant colocalization with sites of active DNA replication. The distribution of kin17 protein is in equilibrium between chromatin-DNA and the nuclear matrix. An increased association with nonchromatin nuclear structure is observed in S-phase cells. We demonstrated here that kin17 protein strongly associates in vivo with DNA fragments containing replication origins in both human HeLa and monkey CV-1 cells. This association was 10-fold higher than that observed with nonorigin control DNA fragments in exponentially growing cells. In addition, the association of kin17 protein to DNA fragments containing replication origins was also analyzed as a function of the cell cycle. High binding of kin17 protein was found at the G1/S border and throughout the S phase and was negligible in both G0 and M phases. Specific monoclonal antibodies against kin17 protein induced a threefold inhibition of in vitro DNA replication of a plasmid containing a minimal replication origin that could be partially restored by the addition of recombinant kin17 protein. Immunoelectron microscopy confirmed the colocalization of kin17 protein with replication proteins like RPA, PCNA, and DNA polymerase α. A two-step chromatographic fractionation of nuclear extracts from HeLa cells revealed that kin17 protein localized in vivo in distinct protein complexes of high molecular weight. We found that kin17 protein purified within an ∼600-kDa protein complex able to support in vitro DNA replication by means of two different biochemical methods designed to isolate replication complexes. In addition, the reduced in vitro DNA replication activity of the multiprotein replication complex after immunodepletion for kin17 protein highlighted for a direct role in DNA replication at the origins.

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