scholarly journals Mandatory coupling of zygotic transcription to DNA replication in early Drosophila embryos

2022 ◽  
Author(s):  
Chun-Yi Cho ◽  
James P. Kemp ◽  
Robert J. Duronio ◽  
Patrick H. O'Farrell
Keyword(s):  
Dna Replication ◽  
Rna Polymerase Ii ◽  
Genome Stability ◽  
Cluster Formation ◽  
Embryonic Cell ◽  
Rna Polymerases ◽  
Replication Forks ◽  
Transcriptional Initiation ◽  
Cell Cycles ◽  
Zygotic Transcription

Collisions between transcribing RNA polymerases and DNA replication forks are disruptive. The threat of collisions is particularly acute during the rapid early embryonic cell cycles of Drosophila when S phase occupies the entirety of interphase. We hypothesized that collision-avoidance mechanisms safeguard the onset of zygotic transcription in these cycles. To explore this hypothesis, we used real-time imaging of transcriptional events at the onset of each interphase. Endogenously tagged RNA polymerase II (RNAPII) abruptly formed clusters before nascent transcripts accumulated, indicating recruitment prior to transcriptional engagement. Injection of inhibitors of DNA replication prevented RNAPII clustering, blocked formation of foci of the pioneer factor Zelda, and largely prevented expression of transcription reporters. Knockdown of Zelda or the histone acetyltransferase CBP prevented RNAPII cluster formation except at the replication-dependent (RD) histone gene locus. We suggest a model in which the passage of replication forks allows Zelda and a distinct pathway at the RD histone locus to reconfigure chromatin to nucleate RNAPII clustering and promote transcriptional initiation. The replication dependency of these events defers initiation of transcription and ensures that RNA polymerases transcribe behind advancing replication forks. The resulting coordination of transcription and replication explains how early embryos circumvent collisions and promote genome stability.

Zygotic expression of the pebble locus is required for cytokinesis during the postblastoderm mitoses of Drosophila

Development ◽  
1992 ◽  
Vol 114 (1) ◽  
pp. 165-171 ◽  
Author(s):  
G. Hime ◽  
R. Saint
Keyword(s):  
Drosophila Melanogaster ◽  
Developmental Regulation ◽  
Single Cells ◽  
Embryonic Cell ◽  
Cell Cycles ◽  
Drosophila Embryogenesis ◽  
Zygotic Transcription ◽  
Homozygous Mutant ◽  
Mitotic Figures

Mutations at the pebble locus of Drosophila melanogaster result in embryonic lethality. Examination of homozygous mutant embryos at the end of embryogenesis revealed the presence of fewer and larger cells which contained enlarged nuclei. Characterization of the embryonic cell cycles using DAPI, propidium iodide, anti-tubulin and anti-spectrin staining showed that the first thirteen rapid syncytial nuclear divisions proceeded normally in pebble mutant embryos. Following cellularization, the postblastoderm nuclear divisions occurred (mitoses 14, 15 and 16), but cytokinesis was never observed. Multinucleate cells and duplicate mitotic figures were seen within single cells at the time of the cycle 15 mitoses. We conclude that zygotic expression of the pebble gene is required for cytokinesis following cellularization during Drosophila embryogenesis. We postulate that developmental regulation of zygotic transcription of the pebble gene is a consequence of the transition from syncytial to cellular mitoses during cycle 14 of embryogenesis.

scholarly journals ING2 controls the progression of DNA replication forks to maintain genome stability

EMBO Reports ◽  
2009 ◽  
Vol 10 (10) ◽  
pp. 1168-1174 ◽  
Author(s):  
Delphine Larrieu ◽  
Damien Ythier ◽  
Romuald Binet ◽  
Christian Brambilla ◽  
Elisabeth Brambilla ◽  
...  
Keyword(s):  
Dna Replication ◽  
Genome Stability ◽  
Replication Forks ◽  
Maintain Genome Stability

scholarly journals Nucleolar Localization and Dynamic Roles of Flap Endonuclease 1 in Ribosomal DNA Replication and Damage Repair

2008 ◽  
Vol 28 (13) ◽  
pp. 4310-4319 ◽  
Author(s):  
Zhigang Guo ◽  
Limin Qian ◽  
Ren Liu ◽  
Huifang Dai ◽  
Mian Zhou ◽  
...  
Keyword(s):  
Dna Replication ◽  
Genome Stability ◽  
Phosphorylation Site ◽  
Replication Forks ◽  
Repair Capacity ◽  
Flap Endonuclease 1 ◽  
Damage Repair ◽  
Dependent Endonuclease ◽  
Cross Links ◽  
Cellular Compartmentalization

ABSTRACT Despite the wealth of information available on the biochemical functions and our recent findings of its roles in genome stability and cancer avoidance of the structure-specific flap endonuclease 1 (FEN1), its cellular compartmentalization and dynamics corresponding to its involvement in various DNA metabolic pathways are not yet elucidated. Several years ago, we demonstrated that FEN1 migrates into the nucleus in response to DNA damage and under certain cell cycle conditions. In the current paper, we found that FEN1 is superaccumulated in the nucleolus and plays a role in the resolution of stalled DNA replication forks formed at the sites of natural replication fork barriers. In response to UV irradiation and upon phosphorylation, FEN1 migrates to nuclear plasma to participate in the resolution of UV cross-links on DNA, most likely employing its concerted action of exonuclease and gap-dependent endonuclease activities. Based on yeast complementation experiments, the mutation of Ser187Asp, mimicking constant phosphorylation, excludes FEN1 from nucleolar accumulation. The replacement of Ser187 by Ala, eliminating the only phosphorylation site, retains FEN1 in nucleoli. Both of the mutations cause UV sensitivity, impair cellular UV damage repair capacity, and decline overall cellular survivorship.

scholarly journals Werner Syndrome Protein and DNA Replication

2018 ◽  
Vol 19 (11) ◽  
pp. 3442 ◽  
Author(s):  
Shibani Mukherjee ◽  
Debapriya Sinha ◽  
Souparno Bhattacharya ◽  
Kalayarasan Srinivasan ◽  
Salim Abdisalaam ◽  
...  
Keyword(s):  
Dna Replication ◽  
Ataxia Telangiectasia ◽  
Replication Fork ◽  
Genome Stability ◽  
Werner Syndrome ◽  
Genotoxic Stress ◽  
Premature Aging ◽  
Replication Forks ◽  
Werner Syndrome Protein ◽  
Syndrome Protein

Werner Syndrome (WS) is an autosomal recessive disorder characterized by the premature development of aging features. Individuals with WS also have a greater predisposition to rare cancers that are mesenchymal in origin. Werner Syndrome Protein (WRN), the protein mutated in WS, is unique among RecQ family proteins in that it possesses exonuclease and 3′ to 5′ helicase activities. WRN forms dynamic sub-complexes with different factors involved in DNA replication, recombination and repair. WRN binding partners either facilitate its DNA metabolic activities or utilize it to execute their specific functions. Furthermore, WRN is phosphorylated by multiple kinases, including Ataxia telangiectasia mutated, Ataxia telangiectasia and Rad3 related, c-Abl, Cyclin-dependent kinase 1 and DNA-dependent protein kinase catalytic subunit, in response to genotoxic stress. These post-translational modifications are critical for WRN to function properly in DNA repair, replication and recombination. Accumulating evidence suggests that WRN plays a crucial role in one or more genome stability maintenance pathways, through which it suppresses cancer and premature aging. Among its many functions, WRN helps in replication fork progression, facilitates the repair of stalled replication forks and DNA double-strand breaks associated with replication forks, and blocks nuclease-mediated excessive processing of replication forks. In this review, we specifically focus on human WRN’s contribution to replication fork processing for maintaining genome stability and suppressing premature aging. Understanding WRN’s molecular role in timely and faithful DNA replication will further advance our understanding of the pathophysiology of WS.

scholarly journals A new role of the Mus81 nuclease for replication completion after fork restart

2019 ◽  
Author(s):  
Benjamin Pardo ◽  
María Moriel-Carretero ◽  
Thibaud Vicat ◽  
Andrés Aguilera ◽  
Philippe Pasero
Keyword(s):  
Dna Replication ◽  
Topoisomerase I ◽  
Genome Stability ◽  
Molecular Level ◽  
Dna Supercoiling ◽  
Replication Forks ◽  
Dna Topoisomerase ◽  
Replication Restart ◽  
Protein Crosslinks

ABSTRACTImpediments to DNA replication threaten genome stability. The homologous recombination (HR) pathway is involved in the restart of blocked replication forks. Here, we used a new method to study at the molecular level the restart of replication in response to DNA topoisomerase I poisoning by camptothecin (CPT). We show that HR-mediated restart at the global genomic level occurs by a BIR-like mechanism that requires Rad52, Rad51 and Pol32. The Mus81 endonuclease, previously proposed to cleave blocked forks, is not required for replication restart in S phase but appears to be essential to resolve fork-associated recombination intermediates in G2/M as a step necessary to complete replication. We confirmed our results using an independent system that allowed us to conclude that this mechanism is independent of the accumulation of DNA supercoiling and DNA-protein crosslinks normally caused by CPT. Thus, we describe here a specific function for Mus81 in the processing of HR-restarted forks required to complete DNA replication.

scholarly journals Spatiotemporal coupling and decoupling of gene transcription with DNA replication origins during embryogenesis in C. elegans

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Ehsan Pourkarimi ◽  
James M Bellush ◽  
Iestyn Whitehouse
Keyword(s):  
Dna Replication ◽  
Developmental Stages ◽  
Embryonic Cell ◽  
Genome Replication ◽  
Replication Origins ◽  
C Elegans ◽  
Cellular Changes ◽  
Zygotic Transcription ◽  
Close Proximity ◽  
Dna Replication Origins

The primary task of developing embryos is genome replication, yet how DNA replication is integrated with the profound cellular changes that occur through development is largely unknown. Using an approach to map DNA replication at high resolution in C. elegans, we show that replication origins are marked with specific histone modifications that define gene enhancers. We demonstrate that the level of enhancer associated modifications scale with the efficiency at which the origin is utilized. By mapping replication origins at different developmental stages, we show that the positions and activity of origins is largely invariant through embryogenesis. Contrary to expectation, we find that replication origins are specified prior to the broad onset of zygotic transcription, yet when transcription initiates it does so in close proximity to the pre-defined replication origins. Transcription and DNA replication origins are correlated, but the association breaks down when embryonic cell division ceases. Collectively, our data indicate that replication origins are fundamental organizers and regulators of gene activity through embryonic development.

scholarly journals TRAIP is a PCNA-binding ubiquitin ligase that protects genome stability after replication stress

2015 ◽  
Vol 212 (1) ◽  
pp. 63-75 ◽  
Author(s):  
Saskia Hoffmann ◽  
Stine Smedegaard ◽  
Kyosuke Nakamura ◽  
Gulnahar B. Mortuza ◽  
Markus Räschle ◽  
...  
Keyword(s):  
Dna Replication ◽  
Ubiquitin Ligase ◽  
Genome Stability ◽  
Replication Stress ◽  
Nuclear Antigen ◽  
Replication Forks ◽  
Chromosomal Dna ◽  
Checkpoint Signaling ◽  
Processivity Factor ◽  
Proliferating Cell

Cellular genomes are highly vulnerable to perturbations to chromosomal DNA replication. Proliferating cell nuclear antigen (PCNA), the processivity factor for DNA replication, plays a central role as a platform for recruitment of genome surveillance and DNA repair factors to replication forks, allowing cells to mitigate the threats to genome stability posed by replication stress. We identify the E3 ubiquitin ligase TRAIP as a new factor at active and stressed replication forks that directly interacts with PCNA via a conserved PCNA-interacting peptide (PIP) box motif. We show that TRAIP promotes ATR-dependent checkpoint signaling in human cells by facilitating the generation of RPA-bound single-stranded DNA regions upon replication stress in a manner that critically requires its E3 ligase activity and is potentiated by the PIP box. Consequently, loss of TRAIP function leads to enhanced chromosomal instability and decreased cell survival after replication stress. These findings establish TRAIP as a PCNA-binding ubiquitin ligase with an important role in protecting genome integrity after obstacles to DNA replication.

scholarly journals SDE2 Integrates into the TIMELESS-TIPIN Complex to Protect Stalled Replication Forks

2020 ◽  
Author(s):  
Julie Rageul ◽  
Jennifer J. Park ◽  
Ping Ping Zeng ◽  
Eun-A Lee ◽  
Jihyeon Yang ◽  
...  
Keyword(s):  
Dna Replication ◽  
Replication Fork ◽  
Genome Stability ◽  
Essential Role ◽  
Replication Stress ◽  
Replication Forks ◽  
Genomic Integrity ◽  
Fork Protection Complex ◽  
Stalled Replication Forks ◽  
Stalled Fork

ABSTRACTProtecting replication fork integrity during DNA replication is essential for maintaining genome stability. Here, we report that SDE2, a PCNA-associated protein, plays a key role in maintaining active replication and counteracting replication stress by regulating the replication fork protection complex (FPC). SDE2 directly interacts with the FPC component TIMELESS (TIM) and enhances TIM stability and its localization to replication forks, thereby aiding the coordination of replisome progression. Like TIM deficiency, knockdown of SDE2 leads to impaired fork progression and stalled fork recovery, along with a failure to activate CHK1 phosphorylation. Moreover, loss of SDE2 or TIM results in an excessive MRE11-dependent degradation of reversed forks. Together, our study uncovers an essential role for SDE2 in maintaining genomic integrity by stabilizing the FPC and describes a new role for TIM in protecting stalled replication forks. We propose that TIM-mediated fork protection may represent a way to cooperate with BRCA-dependent fork stabilization.

scholarly journals Geminin Deficiency Causes a Chk1-dependent G2 Arrest inXenopus

2002 ◽  
Vol 13 (10) ◽  
pp. 3662-3671 ◽  
Author(s):  
Thomas J. McGarry
Keyword(s):  
Dna Replication ◽  
Structural Integrity ◽  
Embryonic Cell ◽  
Embryonic Cells ◽  
Midblastula Transition ◽  
Cell Cycles ◽  
Checkpoint Pathway ◽  
G2 Phase ◽  
Essential Function ◽  
Unusual Phenotype

Geminin is an unstable inhibitor of DNA replication that gets destroyed at the metaphase/anaphase transition. The biological function of geminin has been difficult to determine because it is not homologous to a characterized protein and has pleiotropic effects when overexpressed. Geminin is thought to prevent a second round of initiation during S or G2 phase. In some assays, geminin induces uncommitted embryonic cells to differentiate as neurons. In this study, geminin was eliminated from developing Xenopus embryos by using antisense techniques. Geminin-deficient embryos show a novel and unusual phenotype: they complete the early cleavage divisions normally but arrest in G2 phase immediately after the midblastula transition. The arrest requires Chk1, the effector kinase of the DNA replication/DNA damage checkpoint pathway. The results indicate that geminin has an essential function and that loss of this function prevents entry into mitosis by a Chk1-dependent mechanism. Geminin may be required to maintain the structural integrity of the genome or it may directly down-regulate Chk1 activity. The data also show that during the embryonic cell cycles, rereplication is almost entirely prevented by geminin-independent mechanisms.

scholarly journals Sen1 is required for RNA Polymerase III transcription termination in an R-loop independent manner

2019 ◽  
Author(s):  
Julieta Rivosecchi ◽  
Marc Larochelle ◽  
Camille Teste ◽  
Frédéric Grenier ◽  
Amélie Malapert ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Fission Yeast ◽  
Rna Polymerase Ii ◽  
Genome Stability ◽  
Transcription Termination ◽  
Rna Polymerase Iii ◽  
Rna Polymerases ◽  
Independent Manner ◽  
Polymerase Iii

ABSTRACTR-loop disassembly by the human helicase Senataxin contributes to genome stability and to proper transcription termination at a subset of RNA polymerase II genes. Whether Senataxin-mediated R-loop disassembly also contributes to transcription termination at other classes of genes has remained unclear. Here we show in fission yeast that SenataxinSen1promotes efficient termination of RNA Polymerase III (RNAP3) transcriptionin vivo. In the absence of SenataxinSen1, RNAP3 accumulates downstream of the primary terminator at RNAP3-transcribed genes and produces long exosome-sensitive 3’-extended transcripts. Importantly, neither of these defects was affected by the removal of R-loops. The finding that SenataxinSen1acts as an ancillary factor for RNAP3 transcription terminationin vivochallenges the pre-existing view that RNAP3 terminates transcription autonomously. We propose that Senataxin is a cofactor for transcription termination that has been co-opted by different RNA polymerases in the course of evolution.

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