scholarly journals Replication-dependent histone biosynthesis is coupled to cell-cycle commitment

2021 ◽  
Vol 118 (31) ◽  
pp. e2100178118
Author(s):  
Claire Armstrong ◽  
Sabrina L. Spencer
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Live Cell Imaging ◽  
Current Model ◽  
S Phase ◽  
Strong Increase ◽  
Key Factors ◽  
Histone Gene Expression ◽  
Small Pool ◽  
Histone Synthesis

The current model of replication-dependent (RD) histone biosynthesis posits that RD histone gene expression is coupled to DNA replication, occurring only in S phase of the cell cycle once DNA synthesis has begun. However, several key factors in the RD histone biosynthesis pathway are up-regulated by E2F or phosphorylated by CDK2, suggesting these processes may instead begin much earlier, at the point of cell-cycle commitment. In this study, we use both fixed- and live-cell imaging of human cells to address this question, revealing a hybrid model in which RD histone biosynthesis is first initiated in G1, followed by a strong increase in histone production in S phase of the cell cycle. This suggests a mechanism by which cells that have committed to the cell cycle build up an initial small pool of RD histones to be available for the start of DNA replication, before producing most of the necessary histones required in S phase. Thus, a clear distinction exists at completion of mitosis between cells that are born with the intention of proceeding through the cell cycle and replicating their DNA and cells that have chosen to exit the cell cycle and have no immediate need for histone synthesis.

Control of histone gene expression in Physarum polycephalum. I. Protein synthesis during the cell cycle

1982 ◽  
Vol 57 (1) ◽  
pp. 139-150
Author(s):  
P.N. Schofield ◽  
I.O. Walker
Keyword(s):  
Cell Cycle ◽  
Physarum Polycephalum ◽  
Specific Activity ◽  
S Phase ◽  
Histone Gene ◽  
Synchronous Cultures ◽  
Histone Gene Expression ◽  
Core Histones ◽  
G2 Phase ◽  
Histone Synthesis

Synchronous cultures of Physarum polycephalum were pulsed with [3H]lysine hydrochloride in S and G2 phases of the cell cycle. Plasmodial extracts were separated into nuclear, ribosomal and acid-soluble post-ribosomal cytoplasmic fractions. Core histones could be detected by staining in the nuclear fractions of both S and G2 phases, but were not detected by staining in the cytoplasmic fractions. Newly synthesized histone was present in S-phase nuclei but not in S-phase cytoplasm. The specific activity of newly synthesized histone in G2-phase nuclei decreased by at least 95% compared to S phase and no newly synthesized histone was observed in G2-phase cytoplasmic fractions. Thus histone synthesis is restricted to S phase. There are no free pools of histone in the cytoplasm of Physarum in either S or G2 phases of the cell cycle.

scholarly journals Distinct replication-independent and -dependent phases of histone gene expression during the Physarum cell cycle.

1987 ◽  
Vol 7 (5) ◽  
pp. 1933-1937 ◽  
Author(s):  
J J Carrino ◽  
V Kueng ◽  
R Braun ◽  
T G Laffler
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Dna Replication ◽  
Dna Synthesis ◽  
S Phase ◽  
Histone Gene ◽  
Histone Mrna ◽  
Histone Gene Expression ◽  
Tightly Coupled ◽  
G2 Phase

During the S phase of the cell cycle, histone gene expression and DNA replication are tightly coupled. In mitotically synchronous plasmodia of the myxomycete Physarum polycephalum, which has no G1 phase, histone mRNA synthesis begins in mid-G2 phase. Although histone gene transcription is activated in the absence of significant DNA synthesis, our data demonstrate that histone gene expression became tightly coupled to DNA replication once the S phase began. There was a transition from the replication-independent phase to the replication-dependent phase of histone gene expression. During the first phase, histone mRNA synthesis appears to be under direct cell cycle control; it was not coupled to DNA replication. This allowed a pool of histone mRNA to accumulate in late G2 phase, in anticipation of future demand. The second phase began at the end of mitosis, when the S phase began, and expression became homeostatically coupled to DNA replication. This homeostatic control required continuing protein synthesis, since cycloheximide uncoupled transcription from DNA synthesis. Nuclear run-on assays suggest that in P. polycephalum this coupling occurs at the level of transcription. While histone gene transcription appears to be directly switched on in mid-G2 phase and off at the end of the S phase by cell cycle regulators, only during the S phase was the level of transcription balanced with the rate of DNA synthesis.

Distinct replication-independent and -dependent phases of histone gene expression during the Physarum cell cycle

1987 ◽  
Vol 7 (5) ◽  
pp. 1933-1937
Author(s):  
J J Carrino ◽  
V Kueng ◽  
R Braun ◽  
T G Laffler
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Dna Replication ◽  
Dna Synthesis ◽  
S Phase ◽  
Histone Gene ◽  
Histone Mrna ◽  
Histone Gene Expression ◽  
Tightly Coupled ◽  
G2 Phase

During the S phase of the cell cycle, histone gene expression and DNA replication are tightly coupled. In mitotically synchronous plasmodia of the myxomycete Physarum polycephalum, which has no G1 phase, histone mRNA synthesis begins in mid-G2 phase. Although histone gene transcription is activated in the absence of significant DNA synthesis, our data demonstrate that histone gene expression became tightly coupled to DNA replication once the S phase began. There was a transition from the replication-independent phase to the replication-dependent phase of histone gene expression. During the first phase, histone mRNA synthesis appears to be under direct cell cycle control; it was not coupled to DNA replication. This allowed a pool of histone mRNA to accumulate in late G2 phase, in anticipation of future demand. The second phase began at the end of mitosis, when the S phase began, and expression became homeostatically coupled to DNA replication. This homeostatic control required continuing protein synthesis, since cycloheximide uncoupled transcription from DNA synthesis. Nuclear run-on assays suggest that in P. polycephalum this coupling occurs at the level of transcription. While histone gene transcription appears to be directly switched on in mid-G2 phase and off at the end of the S phase by cell cycle regulators, only during the S phase was the level of transcription balanced with the rate of DNA synthesis.

scholarly journals Regulation of DNA Replication Licensing and Re-Replication by Cdt1

2021 ◽  
Vol 22 (10) ◽  
pp. 5195
Author(s):  
Hui Zhang
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Replication ◽  
Genome Stability ◽  
E3 Ligase ◽  
S Phase ◽  
Replication Initiation ◽  
Nuclear Antigen ◽  
Repair Synthesis ◽  
Ubiquitin E3 Ligase

In eukaryotic cells, DNA replication licensing is precisely regulated to ensure that the initiation of genomic DNA replication in S phase occurs once and only once for each mitotic cell division. A key regulatory mechanism by which DNA re-replication is suppressed is the S phase-dependent proteolysis of Cdt1, an essential replication protein for licensing DNA replication origins by loading the Mcm2-7 replication helicase for DNA duplication in S phase. Cdt1 degradation is mediated by CRL4Cdt2 ubiquitin E3 ligase, which further requires Cdt1 binding to proliferating cell nuclear antigen (PCNA) through a PIP box domain in Cdt1 during DNA synthesis. Recent studies found that Cdt2, the specific subunit of CRL4Cdt2 ubiquitin E3 ligase that targets Cdt1 for degradation, also contains an evolutionarily conserved PIP box-like domain that mediates the interaction with PCNA. These findings suggest that the initiation and elongation of DNA replication or DNA damage-induced repair synthesis provide a novel mechanism by which Cdt1 and CRL4Cdt2 are both recruited onto the trimeric PCNA clamp encircling the replicating DNA strands to promote the interaction between Cdt1 and CRL4Cdt2. The proximity of PCNA-bound Cdt1 to CRL4Cdt2 facilitates the destruction of Cdt1 in response to DNA damage or after DNA replication initiation to prevent DNA re-replication in the cell cycle. CRL4Cdt2 ubiquitin E3 ligase may also regulate the degradation of other PIP box-containing proteins, such as CDK inhibitor p21 and histone methylase Set8, to regulate DNA replication licensing, cell cycle progression, DNA repair, and genome stability by directly interacting with PCNA during DNA replication and repair synthesis.

Cdc6p establishes and maintains a state of replication competence during G1 phase

1997 ◽  
Vol 110 (6) ◽  
pp. 753-763 ◽  
Author(s):  
C.S. Detweiler ◽  
J.J. Li
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Transcriptional Repression ◽  
Prolonged Exposure ◽  
S Phase ◽  
Restrictive Temperature ◽  
Yeast Saccharomyces Cerevisiae ◽  
Important Intermediate ◽  
Initiation Of Dna Replication ◽  
And Function

CDC6 is essential for the initiation of DNA replication in the budding yeast Saccharomyces cerevisiae. Here we examine the timing of Cdc6p expression and function during the cell cycle. Cdc6p is expressed primarily between mitosis and Start. This pattern of expression is due in part to posttranscriptional controls, since it is maintained when CDC6 is driven by a constitutively induced promoter. Transcriptional repression of CDC6 or exposure of cdc6-1(ts) cells to the restrictive temperature at mitosis blocks subsequent S phase, demonstrating that the activity of newly synthesized Cdc6p is required each cell cycle for DNA replication. In contrast, similar perturbations imposed on cells arrested in G(1) before Start have moderate or no effects on DNA replication. This suggests that, between mitosis and Start, Cdc6p functions in an early step of initiation, effectively making cells competent for replication. Prolonged exposure of cdc6-1(ts) cells to the restrictive temperature at the pre-Start arrest eventually does cripple S phase, indicating that Cdc6p also functions to maintain this initiation competence during G(1). The requirement for Cdc6p to establish and maintain initiation competence tightly correlates with the requirement for Cdc6p to establish and maintain the pre-replicative complex at a replication origin, strongly suggesting that the pre-replicative complex is an important intermediate for the initiation of DNA replication. Confining assembly of the complex to G(1) by restricting expression of Cdc6p to this period may be one way of ensuring precisely one round of replication per cell cycle.

Identification of a new set of cell cycle-regulatory genes that regulate S-phase transcription of histone genes in Saccharomyces cerevisiae

1992 ◽  
Vol 12 (11) ◽  
pp. 5249-5259 ◽  
Author(s):  
H Xu ◽  
U J Kim ◽  
T Schuster ◽  
M Grunstein
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Dna Replication ◽  
Gene Transcription ◽  
S Phase ◽  
Histone Gene ◽  
Mrna Synthesis ◽  
Histone Mrna ◽  
Complementation Groups ◽  
Histone Gene Transcription

Histone mRNA synthesis is tightly regulated to S phase of the yeast Saccharomyces cerevisiae cell cycle as a result of transcriptional and posttranscriptional controls. Moreover, histone gene transcription decreases rapidly if DNA replication is inhibited by hydroxyurea or if cells are arrested in G1 by the mating pheromone alpha-factor. To identify the transcriptional controls responsible for cycle-specific histone mRNA synthesis, we have developed a selection for mutations which disrupt this process. Using this approach, we have isolated five mutants (hpc1, hpc2, hpc3, hpc4, and hpc5) in which cell cycle regulation of histone gene transcription is altered. All of these mutations are recessive and belong to separate complementation groups. Of these, only one (hpc1) falls in one of the three complementation groups identified previously by other means (M. A. Osley and D. Lycan, Mol. Cell. Biol. 7:4204-4210, 1987), indicating that at least seven different genes are involved in the cell cycle-specific regulation of histone gene transcription. hpc4 is unique in that derepression occurs only in the presence of hydroxyurea but not alpha-factor, suggesting that at least one of the regulatory factors is specific to histone gene transcription after DNA replication is blocked. One of the hpc mutations (hpc2) suppresses delta insertion mutations in the HIS4 and LYS2 loci. This effect allowed the cloning and sequence analysis of HPC2, which encodes a 67.5-kDa, highly charged basic protein.

scholarly journals Identification of DHX9 as a cell cycle regulated nucleolar recruitment factor for CIZ1

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Urvi Thacker ◽  
Tekle Pauzaite ◽  
James Tollitt ◽  
Maria Twardowska ◽  
Charlotte Harrison ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Cell Cycle Progression ◽  
Zinc Finger Protein ◽  
S Phase ◽  
Cycle Progression ◽  
Inactive X Chromosome ◽  
Functional Characterisation ◽  
Interaction Partners

Abstract CIP1-interacting zinc finger protein 1 (CIZ1) is a nuclear matrix associated protein that facilitates a number of nuclear functions including initiation of DNA replication, epigenetic maintenance and associates with the inactive X-chromosome. Here, to gain more insight into the protein networks that underpin this diverse functionality, molecular panning and mass spectrometry are used to identify protein interaction partners of CIZ1, and CIZ1 replication domain (CIZ1-RD). STRING analysis of CIZ1 interaction partners identified 2 functional clusters: ribosomal subunits and nucleolar proteins including the DEAD box helicases, DHX9, DDX5 and DDX17. DHX9 shares common functions with CIZ1, including interaction with XIST long-non-coding RNA, epigenetic maintenance and regulation of DNA replication. Functional characterisation of the CIZ1-DHX9 complex showed that CIZ1-DHX9 interact in vitro and dynamically colocalise within the nucleolus from early to mid S-phase. CIZ1-DHX9 nucleolar colocalisation is dependent upon RNA polymerase I activity and is abolished by depletion of DHX9. In addition, depletion of DHX9 reduced cell cycle progression from G1 to S-phase in mouse fibroblasts. The data suggest that DHX9-CIZ1 are required for efficient cell cycle progression at the G1/S transition and that nucleolar recruitment is integral to their mechanism of action.

scholarly journals Human Parvovirus B19 Infection Causes Cell Cycle Arrest of Human Erythroid Progenitors at Late S Phase That Favors Viral DNA Replication

2013 ◽  
Vol 87 (23) ◽  
pp. 12766-12775 ◽  
Author(s):  
Yong Luo ◽  
Steve Kleiboeker ◽  
Xuefeng Deng ◽  
Jianming Qiu
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Dna Content ◽  
Parvovirus B19 ◽  
S Phase ◽  
Human Parvovirus B19 ◽  
Viral Dna ◽  
Viral Dna Replication ◽  
Cycle Arrest ◽  
Cellular Dna

Human parvovirus B19 (B19V) infection has a unique tropism to human erythroid progenitor cells (EPCs) in human bone marrow and the fetal liver. It has been reported that both B19V infection and expression of the large nonstructural protein NS1 arrested EPCs at a cell cycle status with a 4 N DNA content, which was previously claimed to be “G2/M arrest.” However, a B19V mutant infectious DNA (M20mTAD2) replicated well in B19V-semipermissive UT7/Epo-S1 cells but did not induce G2/M arrest (S. Lou, Y. Luo, F. Cheng, Q. Huang, W. Shen, S. Kleiboeker, J. F. Tisdale, Z. Liu, and J. Qiu, J. Virol.86:10748–10758, 2012). To further characterize cell cycle arrest during B19V infection of EPCs, we analyzed the cell cycle change using 5-bromo-2′-deoxyuridine (BrdU) pulse-labeling and DAPI (4′,6-diamidino-2-phenylindole) staining, which precisely establishes the cell cycle pattern based on both cellular DNA replication and nuclear DNA content. We found that although both B19V NS1 transduction and infection immediately arrested cells at a status of 4 N DNA content, B19V-infected 4 N cells still incorporated BrdU, indicating active DNA synthesis. Notably, the BrdU incorporation was caused neither by viral DNA replication nor by cellular DNA repair that could be initiated by B19V infection-induced cellular DNA damage. Moreover, several S phase regulators were abundantly expressed and colocalized within the B19V replication centers. More importantly, replication of the B19V wild-type infectious DNA, as well as the M20mTAD2mutant, arrested cells at S phase. Taken together, our results confirmed that B19V infection triggers late S phase arrest, which presumably provides cellular S phase factors for viral DNA replication.

scholarly journals A Role for Cdk2 Kinase in Negatively Regulating DNA Replication during S Phase of the Cell Cycle

1997 ◽  
Vol 137 (1) ◽  
pp. 183-192 ◽  
Author(s):  
Xuequn Helen Hua ◽  
Hong Yan ◽  
John Newport
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Cyclin E ◽  
Embryonic Cell ◽  
S Phase ◽  
Negative Regulation ◽  
Sperm Chromatin ◽  
High Concentration ◽  
Low Concentrations ◽  
Embryonic Cell Cycle

Using cell-free extracts made from Xenopus eggs, we show that cdk2-cyclin E and A kinases play an important role in negatively regulating DNA replication. Specifically, we demonstrate that the cdk2 kinase concentration surrounding chromatin in extracts increases 200-fold once the chromatin is assembled into nuclei. Further, we find that if the cdk2–cyclin E or A concentration in egg cytosol is increased 16-fold before the addition of sperm chromatin, the chromatin fails to initiate DNA replication once assembled into nuclei. This demonstrates that cdk2–cyclin E or A can negatively regulate DNA replication. With respect to how this negative regulation occurs, we show that high levels of cdk2–cyclin E do not block the association of the protein complex ORC with sperm chromatin but do prevent association of MCM3, a protein essential for replication. Importantly, we find that MCM3 that is prebound to chromatin does not dissociate when cdk2– cyclin E levels are increased. Taken together our results strongly suggest that during the embryonic cell cycle, the low concentrations of cdk2–cyclin E present in the cytosol after mitosis and before nuclear formation allow proteins essential for potentiating DNA replication to bind to chromatin, and that the high concentration of cdk2–cyclin E within nuclei prevents MCM from reassociating with chromatin after replication. This situation could serve, in part, to limit DNA replication to a single round per cell cycle.

Reducing MCM Loading Causes Chromosomal Aneuploidy.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3349-3349
Author(s):  
Stephen J. Orr ◽  
Terry Gaymes ◽  
Rong Wang ◽  
Barbara Czepulkowski ◽  
Darius Ladon ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
T Cells ◽  
Dna Replication ◽  
Genomic Instability ◽  
Chromosomal Abnormalities ◽  
S Phase ◽  
Human T Cells ◽  
Dna Damage Responses ◽  
Mcm Proteins

Abstract Normal DNA replication must be accurate and occur only once per cell cycle. Sites of DNA replication are specified by binding the origin recognition complex, that includes minichromosome maintenance (MCM) proteins. Paradoxically, in higher eukaryotes MCM proteins are present in >20 fold excess of that required for DNA replication. They are also downregulated by elevated expression of proteins such as cyclin E that occurs in cancers, including AML and breast cancer. We investigated why human cells need “excess” MCM proteins and whether the reduction of MCM protein levels might contribute to a malignant phenotype. We determined the consequences of reducing the levels of MCM proteins in primary human T cells in which cell cycle controls and DNA damage responses are normal. Mass spectrometry sequencing of chromatin/nuclear matrix-bound proteins and western blotting identified that Mcm7 is not present in quiescent, normal primary human T cells. Mcm7 is induced in mid G1after the G0→G1 commitment point, the point beyond which T cells are committed to entering the cell cycle. Reduction of Mcm7 with siRNA to <5% of normal during G0→G1→S-phase reduces chromatin-binding of each of the MCM proteins that form the DNA helicase. However, these cells still enter S-phase and replicate DNA. Reducing MCM levels by titrating siRNA causes dose-dependent DNA-damage responses involving activation of ATR & ATM and Chk1 & Chk2. However, cells depleted of Mcm7 do not undergo apoptosis, rather reducing MCM levels even by 50% causes gross non-clonal chromosomal abnormalities normally found in genomic instability syndromes. M-FISH identified chromosome translocations, as well as loss and gain of individual chromosomes, which can occur individually or together in the same cell. Reducing MCM levels also causes misrepair by non-homologous end joining (NHEJ), and both NHEJ and homologous recombination (HR) are necessary for chromosomal abnormalities to occur. Therefore, “excess” MCM proteins that are present in a normal, proliferating cell are necessary for maintaining genome stability and reduction of MCM loading onto DNA that occurs in cancers is sufficient to cause genomic instability.

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