scholarly journals Autophosphorylation of Archaeal Cdc6 Homologues Is Regulated by DNA

2001 ◽  
Vol 183 (18) ◽  
pp. 5459-5464 ◽  
Author(s):  
Beatrice Grabowski ◽  
Zvi Kelman
Keyword(s):  
Dna Replication ◽  
Essential Role ◽  
Biochemical Properties ◽  
Replication Initiation ◽  
Winged Helix ◽  
Initiator Protein ◽  
C Terminus ◽  
Eukaryotic Dna ◽  
Initiation Of Dna Replication ◽  
Winged Helix Domain

ABSTRACT The initiator protein Cdc6 (Cdc18 in fission yeast) plays an essential role in the initiation of eukaryotic DNA replication. In yeast the protein is expressed before initiation of DNA replication and is thought to be essential for loading of the helicase onto origin DNA. The biochemical properties of the protein, however, are largely unknown. Using three archaeal homologues of Cdc6, it was found that the proteins are autophosphorylated on Ser residues. The winged-helix domain at the C terminus of Cdc6 interacts with DNA, which apparently regulates the autophosphorylation reaction. Yeast Cdc18 was also found to autophosphorylate, suggesting that this function of Cdc6 may play a widely conserved and essential role in replication initiation.

A Novel Fluorescence-Based Screen for Inhibitors of the Initiation of DNA Replication in Bacteria

2019 ◽  
Vol 16 (3) ◽  
pp. 272-277 ◽  
Author(s):  
Rasmus N. Klitgaard ◽  
Anders Løbner-Olesen
Keyword(s):  
Dna Replication ◽  
High Throughput ◽  
Cyclic Peptide ◽  
Replication Initiation ◽  
False Positive Result ◽  
Over Expression ◽  
Initiator Protein ◽  
Dna Replication Initiation ◽  
Cellular Processes ◽  
Initiation Of Dna Replication

Background:One of many strategies to overcome antibiotic resistance is the discovery of compounds targeting cellular processes, which have not yet been exploited.Materials and Methods:Using various genetic tools, we constructed a novel high throughput, cellbased, fluorescence screen for inhibitors of chromosome replication initiation in bacteria.Results:The screen was validated by expression of an intra-cellular cyclic peptide interfering with the initiator protein DnaA and by over-expression of the negative initiation regulator SeqA. We also demonstrated that neither tetracycline nor ciprofloxacin triggers a false positive result. Finally, 400 extracts isolated mainly from filamentous actinomycetes were subjected to the screen.Conclusion:We concluded that the presented screen is applicable for identifying putative inhibitors of DNA replication initiation in a high throughput setup.

scholarly journals Multiple pathways can bypass the essential role of fission yeast Hsk1 kinase in DNA replication initiation

2011 ◽  
Vol 195 (3) ◽  
pp. 387-401 ◽  
Author(s):  
Seiji Matsumoto ◽  
Motoshi Hayano ◽  
Yutaka Kanoh ◽  
Hisao Masai
Keyword(s):  
Dna Replication ◽  
Fission Yeast ◽  
Replication Fork ◽  
Essential Role ◽  
Replication Initiation ◽  
Origin Firing ◽  
Physiological Conditions ◽  
Dna Replication Initiation ◽  
Initiation Of Dna Replication

Cdc7/Hsk1 is a conserved kinase required for initiation of DNA replication that potentially regulates timing and locations of replication origin firing. Here, we show that viability of fission yeast hsk1Δ cells can be restored by loss of mrc1, which is required for maintenance of replication fork integrity, by cds1Δ, or by a checkpoint-deficient mutant of mrc1. In these mutants, normally inactive origins are activated in the presence of hydroxyurea and binding of Cdc45 to MCM is stimulated. mrc1Δ bypasses hsk1Δ more efficiently because of its checkpoint-independent inhibitory functions. Unexpectedly, hsk1Δ is viable at 37°C. More DNA is synthesized, and some dormant origins fire in the presence of hydroxyurea at 37°C. Furthermore, hsk1Δ bypass strains grow poorly at 25°C compared with higher temperatures. Our results show that Hsk1 functions for DNA replication can be bypassed by different genetic backgrounds as well as under varied physiological conditions, providing additional evidence for plasticity of the replication program in eukaryotes.

scholarly journals Girdin: an essential component of pre-replicative complex in human cells

2019 ◽  
Author(s):  
Lihong Wu ◽  
Yu Hua ◽  
Feiran Chang ◽  
Daochun Kong
Keyword(s):  
Dna Replication ◽  
Essential Role ◽  
Human Cells ◽  
Replication Initiation ◽  
The Other ◽  
Initiation Factor ◽  
Replication Origins ◽  
Initiation Factors ◽  
Initiation Of Dna Replication ◽  
Dna Replication Origins

AbstractA central event in the initiation of DNA replication in eukaryotes is the assembly of pre-replicative complex (pre-RC) on specific chromatin sites known as DNA replication origins. The pre-RC assembly process differs between budding and fission yeasts. In fission yeast, Sap1 directly participates in pre-RC assembly, together with the four initiation factors: ORC, Cdc18/Cdc6, Cdt1, and MCM. In metazoans, the nature of DNA replication origins is not defined and the mechanism of pre-RC assembly remains incompletely known. In this study, Girdin was identified as an essential replication initiation factor in human cells. Similar to the activity of Sap1, human Girdin binds to DNA origins, interacts with ORC, and is required for pre-RC assembly due to its essential role in recruitment of Cdc6 to DNA origins. Thus, DNA origins in human or metazoans are defined as including two elements, one bound by ORC and the other bound by Girdin.

scholarly journals Polar Remodeling and Histidine Kinase Activation, Which Is Essential for Caulobacter Cell Cycle Progression, Are Dependent on DNA Replication Initiation

2010 ◽  
Vol 192 (15) ◽  
pp. 3893-3902 ◽  
Author(s):  
Antonio A. Iniesta ◽  
Nathan J. Hillson ◽  
Lucy Shapiro
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Histidine Kinase ◽  
Cell Cycle Progression ◽  
Caulobacter Crescentus ◽  
Replication Initiation ◽  
Cycle Progression ◽  
Dna Replication Initiation ◽  
Flagellar Biosynthesis ◽  
Initiation Of Dna Replication

ABSTRACT Caulobacter crescentus initiates a single round of DNA replication during each cell cycle. Following the initiation of DNA replication, the essential CckA histidine kinase is activated by phosphorylation, which (via the ChpT phosphotransferase) enables the phosphorylation and activation of the CtrA global regulator. CtrA∼P then blocks the reinitiation of replication while regulating the transcription of a large number of cell cycle-controlled genes. It has been shown that DNA replication serves as a checkpoint for flagellar biosynthesis and cell division and that this checkpoint is mediated by the availability of active CtrA. Because CckA∼P promotes the activation of CtrA, we addressed the question of what controls the temporal activation of CckA. We found that the initiation of DNA replication is a prerequisite for remodeling the new cell pole, which includes the localization of the DivL protein kinase to that pole and, consequently, the localization, autophosphorylation, and activation of CckA at that pole. Thus, CckA activation is dependent on polar remodeling and a DNA replication initiation checkpoint that is tightly integrated with the polar phospho-signaling cascade governing cell cycle progression.

scholarly journals Control of Initiation of Sporulation by Replication Initiation Genes in Bacillus subtilis

2000 ◽  
Vol 182 (10) ◽  
pp. 2989-2991 ◽  
Author(s):  
Katherine P. Lemon ◽  
Iren Kurtser ◽  
Judy Wu ◽  
Alan D. Grossman
Keyword(s):  
Bacillus Subtilis ◽  
Dna Replication ◽  
Replication Initiation ◽  
Spore Formation ◽  
Temperature Sensitive ◽  
Temperature Sensitive Mutation ◽  
Initiation Of Dna Replication ◽  
Replication Initiation Proteins

ABSTRACT Initiation of spore formation in Bacillus subtilisappears to depend on initiation of DNA replication. This regulation was first identified using a temperature-sensitive mutation indnaB. We found that mutations in the replication initiation genes dnaA and dnaD also inhibit sporulation, indicating that inhibition of sporulation is triggered by general defects in the function of replication initiation proteins.

Initiation of polyoma virus origin-dependent DNA replication through STAT5 activation by human granulocyte-macrophage colony-stimulating factor

Blood ◽  
2001 ◽  
Vol 97 (5) ◽  
pp. 1266-1273 ◽  
Author(s):  
Sumiko Watanabe ◽  
Rong Zeng ◽  
Yutaka Aoki ◽  
Tohru Itoh ◽  
Ken-ichi Arai
Keyword(s):  
Dna Replication ◽  
Transcriptional Activation ◽  
Tandem Repeats ◽  
T Antigen ◽  
Replication Initiation ◽  
Macrophage Colony Stimulating Factor ◽  
Initiation Of Dna Replication ◽  
Macrophage Colony ◽  
Stimulating Factor

Several lines of evidence indicate that transcriptional activation is coupled with DNA replication initiation, but the nature of initiation of DNA replication in mammalian cells is unclear. Polyoma virus replicon is an excellent system to analyze the initiation of DNA replication in murine cells because its replication requires an enhancer, and all components of replication machinery, except for DNA helicase large T antigen, are supplied by host cells. This system was used to examine the role of signal transducer and activator of transcription (STAT5) in replication initiation of polyoma replicon in the mouse lymphoid cell line BA/F3. The plasmid with tandem repeats of consensus STAT5 binding sites followed by polyoma replication origin was replicated by stimulation with human granulocyte-macrophage colony-stimulating factor (hGM-CSF) in the presence of polyoma large T antigen in BA/F3 cells. Mutation analysis of the hGM-CSF receptor β subunit revealed that only the box1 region is essential, and the C-terminal tyrosine residues are dispensable for the activity. Addition of the tyrosine kinase inhibitor genistein suppressed this replication without affecting transcriptional activation of STAT5. Because deletion analysis of STAT5 indicates the importance of the C-terminal transcriptional activation domain of STAT5 for the initiation of replication, the role of this region in the activation of replication was examined with a GAL4–STAT5 fusion protein. GAL4–STAT5 activated replication of the plasmid containing tandem repeats of GAL4 binding sites and polyoma replication origin in BA/F3 cells. Mutation analysis of GAL4–STAT5 indicated that multiple serine residues coordinately have a role in activating replication. This is the first direct evidence indicating the potential involvement of STAT5 in replication.

scholarly journals The role of DDK and Treslin-MTBP in coordinating replication licensing and pre-Initiation Complex formation

2021 ◽  
Author(s):  
Ilaria Volpi ◽  
Peter J. Gillespie ◽  
Gaganmeet Singh Chadha ◽  
J. Julian Blow
Keyword(s):  
Dna Replication ◽  
Complex Formation ◽  
S Phase ◽  
Replication Initiation ◽  
Initiation Complex ◽  
Crucial Step ◽  
Egg Extract ◽  
Initiation Of Dna Replication ◽  
Rate Limiting

AbstractTreslin/Ticrr is required for the initiation of DNA replication and binds to MTBP (Mdm2 Binding Protein). Here we show that in Xenopus egg extract, MTBP forms an elongated tetramer with Treslin containing two molecules of each protein. Immunodepletion and add-back experiments show that Treslin-MTBP is rate-limiting for replication initiation. It is recruited onto chromatin before S phase starts and recruitment continues during S phase. We show that DDK activity both increases and strengthens the interaction of Treslin-MTBP with licensed chromatin. We also show that DDK activity cooperates with CDK activity to drive the interaction of Treslin-MTBP with TopBP1 which is a regulated crucial step in pre-Initiation Complex formation. These results suggest how DDK works together with CDKs to regulate Treslin-MTBP and plays a crucial in selecting which origins will undergo initiation.

scholarly journals The role of DDK and Treslin–MTBP in coordinating replication licensing and pre-initiation complex formation

Open Biology ◽  
2021 ◽  
Vol 11 (10) ◽  
Author(s):  
Ilaria Volpi ◽  
Peter J. Gillespie ◽  
Gaganmeet Singh Chadha ◽  
J. Julian Blow
Keyword(s):  
Dna Replication ◽  
Complex Formation ◽  
S Phase ◽  
Replication Initiation ◽  
Initiation Complex ◽  
Crucial Step ◽  
Egg Extract ◽  
Initiation Of Dna Replication ◽  
Rate Limiting

Treslin/Ticrr is required for the initiation of DNA replication and binds to MTBP (Mdm2 Binding Protein). Here, we show that in Xenopus egg extract, MTBP forms an elongated tetramer with Treslin containing two molecules of each protein. Immunodepletion and add-back experiments show that Treslin–MTBP is rate limiting for replication initiation. It is recruited onto chromatin before S phase starts and recruitment continues during S phase. We show that DDK activity both increases and strengthens the interaction of Treslin–MTBP with licensed chromatin. We also show that DDK activity cooperates with CDK activity to drive the interaction of Treslin–MTBP with TopBP1 which is a regulated crucial step in pre-initiation complex formation. These results suggest how DDK works together with CDKs to regulate Treslin–MTBP and plays a crucial in selecting which origins will undergo initiation.

scholarly journals CDC45, a novel yeast gene that functions with the origin recognition complex and Mcm proteins in initiation of DNA replication.

1997 ◽  
Vol 17 (2) ◽  
pp. 553-563 ◽  
Author(s):  
L Zou ◽  
J Mitchell ◽  
B Stillman
Keyword(s):  
Dna Replication ◽  
Origin Recognition Complex ◽  
Genetic Interaction ◽  
Sequence Similarity ◽  
Replication Initiation ◽  
Nonpermissive Temperature ◽  
Functional Connection ◽  
Initiation Of Dna Replication ◽  
Mcm Proteins ◽  
Origin Recognition

The CDC45 gene of Saccharomyces cerevisiae was isolated by complementation of the cold-sensitive cdc45-1 mutant and shown to be essential for cell viability. Although CDC45 genetically interacts with a group of MCM genes (CDC46, CDC47, and CDC54), the predicted sequence of its protein product reveals no significant sequence similarity to any known Mcm family member. Further genetic characterization of the cdc45-1 mutant demonstrated that it is synthetically lethal with orc2-1, mcm2-1, and mcm3-1. These results not only reveal a functional connection between the origin recognition complex (ORC) and Cdc45p but also extend the CDC45-MCM genetic interaction to all known MCM family members that were shown to be involved in replication initiation. Initiation of DNA replication in cdc45-1 cells was defective, causing a delayed entry into S phase at the nonpermissive temperature, as well as a high plasmid loss rate which could be suppressed by tandem copies of replication origins. Furthermore, two-dimensional gels directly showed that chromosomal origins fired less frequently in cdc45-1 cells at the nonpermissive temperature. These findings suggest that Cdc45p, ORC, and Mcm proteins act in concert for replication initiation throughout the genome.

scholarly journals Control of DNA Replication Initiation by Ubiquitin

Cells ◽  
2018 ◽  
Vol 7 (10) ◽  
pp. 146 ◽  
Author(s):  
Esperanza Hernández-Carralero ◽  
Elisa Cabrera ◽  
Ignacio Alonso-de Vega ◽  
Santiago Hernández-Pérez ◽  
Veronique Smits ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Replication Initiation ◽  
Post Translational Modifications ◽  
Precise Control ◽  
Dna Replication Initiation ◽  
Fundamental Part ◽  
Initiation Of Dna Replication ◽  
The Right ◽  
Hydrolysis Of

Eukaryotic cells divide by accomplishing a program of events in which the replication of the genome is a fundamental part. To ensure all cells have an accurate copy of the genome, DNA replication occurs only once per cell cycle and is controlled by numerous pathways. A key step in this process is the initiation of DNA replication in which certain regions of DNA are marked as competent to replicate. Moreover, initiation of DNA replication needs to be coordinated with other cell cycle processes. At the molecular level, initiation of DNA replication relies, among other mechanisms, upon post-translational modifications, including the conjugation and hydrolysis of ubiquitin. An example is the precise control of the levels of the DNA replication initiation protein Cdt1 and its inhibitor Geminin by ubiquitin-mediated proteasomal degradation. This control ensures that DNA replication occurs with the right timing during the cell cycle, thereby avoiding re-replication events. Here, we review the events that involve ubiquitin signalling during DNA replication initiation, and how they are linked to human disease.

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