Localization of Cdc7 Protein Kinase During DNA Replication in Saccharomyces cerevisiae
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ABSTRACT:
PROVIDER: PRJNA407733 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR6048333.fastq.gz | Fastqsanger.gz | |||
| SRR6048334.fastq.gz | Fastqsanger.gz | |||
| SRR6048335.fastq.gz | Fastqsanger.gz | |||
| SRR6048336.fastq.gz | Fastqsanger.gz |
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Project description:Cdc7 kinase is known to initiate DNA replication, but it is unknown where Cdc7 is found within the genome. We modified the Calling Cards method that uses the Ty5 retrotransposon to investigate Cdc7 binding in the genome. The Ty5 retrotransposon is inserted into the genome by DNA transcription factors or replication factors binding within the genome. We find that Cdc7 inserts Ty5 transposons throughout chromosomes and furthermore creates more Ty5 insertions into regions of DNA that are known to replicate early. Cdc7 does not solely integrate Ty5 at origins or replication, but rather throughout the genome.
2017-09-18 | GSE103943 | GEO
Project description:In eukaryotes, CDC7 kinase is crucial for DNA replication initiation and has been involved in fork processing and replication stress response. Human CDC7 requires the binding of either one of two regulatory subunits, DBF4 and DRF1, for its activity. However, it is unclear whether the two regulatory subunits target CDC7 to a specific set of substrates, thus having different biological functions, or if they act redundantly. Using genome editing technology, we generated an isogenic set of cell lines deficient in either one of the two CDC7-activating subunits: these cells are viable but present signs of genomic instability, indicating that both DBF4 and DRF1 can independently support CDC7 for bulk DNA replication. Nonetheless, DBF4-deficient cells show altered replication efficiency, including partial deficiency in MCM helicase phosphorylation and alterations in the replication timing of discrete genomic regions. Notably, we find that CDC7 function at replication forks is entirely dependent on DBF4 and not DRF1. Thus, DBF4 is the primary regulator of CDC7 activity, likely mediating most of its functions in unperturbed DNA replication and during replication fork processing upon replication interference.
2024-03-28 | GSE248981 | GEO
Project description:We demonstrate that the Cdc7/Dbf4-dependent histone modification, H3 threonine 45 phosphorylation (H3T45p), is specifically enriched at origins of replication, and highly transcribed genes involved in protein synthesis and glycolysis. Furthermore, we show that H3T45p also mediates polymerase recruitment and expression of these genes.
2022-01-09 | GSE43737 | GEO
Project description:DBF4, not DRF1, is the crucial regulator of CDC7 kinase at replication forks.
| PRJNA1046531 | ENA
Project description:Centromeres play several important roles in ensuring proper chromosome segregation. Not only do they promote kinetochore assembly for microtubule attachment, but they also support robust sister chromatid cohesion at pericentromeres and facilitate replication of centromeric DNA early in S phase. However, it is still elusive how centromeres orchestrate all these functions at the same site. Here we show that the budding yeast Dbf4-dependent kinase (DDK) accumulates at kinetochores in telophase, facilitated by the Ctf19 kinetochore complex. This promptly recruits Sld3-Sld7 replication initiator proteins to pericentromeric replication origins so that they initiate replication early in S phase. Furthermore DDK at kinetochores independently recruits the Scc2-Scc4 cohesin loader to centromeres in G1 phase. This enhances cohesin loading and facilitates robust pericentromeric cohesion in S phase. Thus, we have found the central mechanism by which kinetochores orchestrate early S phase DNA replication and robust sister chromatid cohesion at microtubule attachment sites. Measurement of genome replication time for various S. cerevisiae strains. For each strain two samples were analysed: a replicating sample (from S phase) and a non-replicating sample (from G2 phase).
2013-06-06 | E-GEOD-41982 | biostudies-arrayexpress
Project description:CDC7 is a serine-threonine kinase, which has an important role in the activation of replication origins in the S-phase. CDC7 is also a potential molecular target for the development of anticancer drugs. Multiple studies have shown that different cell lines have a wide range of sensitivity to CDC7 inhibition. However, how the manner in which cells respond to prolonged CDC7 inhibition has yet to be investigated. Hence, we performed an RNA-sequencing experiment to investigate variation in gene expression in cells undergoing prolonged CDC7 inhibition.
2024-05-14 | E-MTAB-13179 | biostudies-arrayexpress
Project description:Entry of mammalian cells into DNA synthesis (S-phase) represents a key event in cell division. According to the current cell cycle models, the Cdc7 kinase represents the essential and rate-limiting trigger of DNA replication, acting together with the cyclin-dependent kinase Cdk2. Using chemical genetic systems which allow an acute shutdown of Cdc7 in in vitro cultured cells as well as in vivo in a living mouse, that Cdc7 is dispensable for cell division of many different cell types. Proteomic and phosphoproteomic analysis were performed to elucidate the compensating mechanism. We demonstrate that Cdk1-cyclin B, a kinase currently thought to act exclusively during mitosis, is also active during G1/S transition both in cycling cells and in cells exiting quiescence. We show that Cdc7 and Cdk1-cyclin B play redundant roles during G1/S transition, and at least one of these kinases is needed to allow S-phase entry. These observations revise our fundamental understanding of cell cycle progression by demonstrating that Cdk1-cyclin B physiologically regulates two distinct transitions during cell division cycle, while Cdc7 plays a redundant function in DNA replication. In TMT1, to analyze the expression of Cdc7 and its partners upon auxin-mediated induction of Cdc7 degradation, we performed a TMT analysis of ES cells treated with IAA for 24 hours. In TMT2, to analyze the phosphorylation of Cdc7 targets as a function of time upon degradation of the kinase we performed the time-resolved TMT analysis of ES cells subjected to Cdc7 degradation for three different amounts of time. In TMT3, to determine the ability of Cdk1 to phosphorylate Mcm2, we analyzed the phosphoproteome of mES cells and focused on the phosphorylation of Mcm2 upon Cdk1 inhibition.
2022-03-11 | PXD025625 | Pride
Project description:Centromeres play several important roles in ensuring proper chromosome segregation. Not only do they promote kinetochore assembly for microtubule attachment, but they also support robust sister chromatid cohesion at pericentromeres and facilitate replication of centromeric DNA early in S phase. However, it is still elusive how centromeres orchestrate all these functions at the same site. Here we show that the budding yeast Dbf4-dependent kinase (DDK) accumulates at kinetochores in telophase, facilitated by the Ctf19 kinetochore complex. This promptly recruits Sld3-Sld7 replication initiator proteins to pericentromeric replication origins so that they initiate replication early in S phase. Furthermore DDK at kinetochores independently recruits the Scc2-Scc4 cohesin loader to centromeres in G1 phase. This enhances cohesin loading and facilitates robust pericentromeric cohesion in S phase. Thus, we have found the central mechanism by which kinetochores orchestrate early S phase DNA replication and robust sister chromatid cohesion at microtubule attachment sites.
2013-06-06 | GSE41982 | GEO
Project description:This SuperSeries is composed of the following subset Series: GSE9233: Genome-wide localization of Ino80 and Arp5 on chromosome in Saccharomyces cerevisiae GSE9420: Genome-wide localization of Ino80 on chromosome in Saccharomyces cerevisiae Keywords: SuperSeries Refer to individual Series
2008-04-10 | E-GEOD-9421 | biostudies-arrayexpress
Project description:DDK, a conserved serine-threonine protein kinase composed of a regulatory subunit, Dbf4, and a catalytic subunit, Cdc7, is essential for DNA replication initiation during S phase of the cell cycle through MCM2-7 helicase phosphorylation. The biological significance of DDK is well characterized, but the full mechanism of how DDK associates with substrates remains unclear. Cdc7 is bound to chromatin in the Saccharomyces cerevisiae genome throughout the cell cycle, but there is little empirical evidence as to specific Cdc7 binding locations. Using biochemical and genetic techniques, this study investigated the specific localization of Cdc7 on chromatin. The Calling Cards method, using Ty5 retrotransposons as a marker for DNA-protein binding, suggests Cdc7 kinase is preferentially bound to genomic DNA known to replicate early in S phase, including centromeres and origins of replication. We also discovered Cdc7 binding throughout the genome, which may be necessary to initiate other cellular processes, including meiotic recombination and translesion synthesis. A kinase dead Cdc7 point mutation increases the Ty5 retrotransposon integration efficiency and a 55-amino acid C-terminal truncation of Cdc7, unable to bind Dbf4, reduces Cdc7 binding suggesting a requirement for Dbf4 to stabilize Cdc7 on chromatin during S phase. Chromatin immunoprecipitation demonstrates that Cdc7 binding near specific origins changes during S phase. Our results suggest a model where Cdc7 is loosely bound to chromatin during G1 At the G1/S transition, Cdc7 binding to chromatin is increased and stabilized, preferentially at sites that may become origins, in order to carry out a variety of cellular processes.
| S-EPMC5677158 | biostudies-literature