Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Proteomic profiling reveals distinct phases to the restoration of chromatin following DNA replication

ABSTRACT: Chromatin organization must be maintained during cell proliferation to preserve cellular identity and genome integrity. However, DNA replication results in transient displacement of DNA bound proteins, and it is unclear how they regain access to newly replicated DNA. Using quantitative MS-based proteomics coupled to isolation of proteins on nascent DNA (iPOND), we provide time resolved binding kinetics for thousands of proteins behind replisomes in the lung fibroblast cell line TIG-3 (JCRB0506, JCRB Cell Bank). Performing hierarchical clustering, we identified four groups of protein behaviour: Transient enriched on nascent, restored within 11min, peaking in G2/M, and delayed restored. Overall, our data show that most proteins (85%) regain access within the first 2h after the passage of the fork. Only a small percentage of proteins is restored after 2h post-replication, with 5.3 % of factors peaking in G2/M and 9.1% factors not restored before the next G1 phase. We provide evidence that DNA replication not only disrupts but also promotes recruitment of transcription factors and chromatin remodelers, providing a significant advance in understanding how DNA replication could contribute to programmed changes of cell memory.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Vanesa Alvarez

LAB HEAD: Constance Alabert

PROVIDER: PXD036236 | Pride | 2023-02-02

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	Data_information.xlsx	Xlsx
	Homo_sapiens_SpTr_canonical_20190902.fasta	Fasta
	IP_combined.zip	Other
	Input_combined.zip	Other
	Input_mqpar.xml	Xml

Items per page:

1 - 5 of 206

Similar Datasets

Proteomic profiling reveals distinct phases to the restoration of chromatin following DNA replication

Project description:Chromatin organization must be maintained during cell proliferation to preserve cellular identity and genome integrity. However, DNA replication results in transient displacement of DNA bound proteins, and it is unclear how they regain access to newly replicated DNA. Using quantitative MS-based proteomics coupled to Nascent Chromatin Capture, we provide time resolved binding kinetics for thousands of proteins behind replisomes of mid-S replicated regions. This shows that most proteins regain access within the first 15 minutes after the passage of the fork. In contrast, 25% of the identified proteins do not, and this delay cannot be inferred from their known function, physicochemical properties, or nuclear abundance. A sample treated with nocodazole to block the cell cycle progression. The comparison between a normal cell cycle progression and a nocodazole treated sample shows that the reassociation onto chromatin of proteins with a delayed restoration can be regulated by the cell cycle progression and/or the post-replication time. In addition, we provide evidence that DNA replication not only disrupts but also promotes recruitment of transcription factors and chromatin remodellers, providing a significant advance in understanding how DNA replication could contribute to programmed changes of cell memory. Data available in this entry are related with the supplementary table 5 from our manuscript.

2023-02-02 | PXD038113 | Pride

Proteomic profiling reveals distinct phases to the restoration of chromatin following DNA replication

Project description:Chromatin organization must be maintained during cell proliferation to preserve cellular identity and genome integrity. However, DNA replication results in transient displacement of DNA bound proteins, and it is unclear how they regain access to newly replicated DNA. Using quantitative MS-based proteomics coupled to Nascent Chromatin Capture, we provide time resolved binding kinetics for thousands of proteins behind replisomes within euchromatin and heterochromatin in human cells. This shows that most proteins regain access within the first 15 minutes after the passage of the fork. In contrast, 30% of the identified proteins do not, and this delay cannot be inferred from their known function, physicochemical properties, or nuclear abundance. Instead, differential chromatin organization affect their reassociation. We provide evidence that DNA replication not only disrupts but also promotes recruitment of transcription factors and chromatin remodellers, providing a significant advance in understanding how DNA replication could contribute to programmed changes of cell memory.We include here a reference to link the data with our manuscript:Tables SUPP1 and SUPP2:NCC data: PT7988 (Bio-rep1), PT8242 (Bio-rep2), PT8243 (Bio-rep3). 1-24 (24 high-pH RPLC fractions from TMT labeled peptides)Nuclear fraction data: PT7988 (Bio-rep1), PT8242 (Bio-rep2), PT8243 (Bio-rep3). 25-48 (24 high-pH RPLC fractions from TMT labeled peptides)Tables SUPP3-5:NCC data: PT9825 (Bio-rep1), PT9825 (Bio-rep2), PT9825 (Bio-rep3). (11-34, 35-58, 59-82) (24 high-pH RPLC fractions from TMT labeled peptides)Nuclear fraction data: PT9825 (Bio-rep1), PT9825 (Bio-rep2), PT9825 (Bio-rep3). (83-106, 107-130, 131-154). (24 high-pH RPLC fractions from TMT labeled peptides)

2023-02-02 | PXD035489 | Pride

Proteomic profiling reveals distinct phases to the restoration of chromatin following DNA replication

Project description:Chromatin organization must be maintained during cell proliferation to preserve cellular identity and genome integrity. However, DNA replication results in transient displacement of DNA bound proteins, and it is unclear how they regain access to newly replicated DNA. Using quantitative MS-based proteomics coupled to Nascent Chromatin Capture, we provide time resolved binding kinetics for thousands of proteins behind replisomes of mid-S replicated regions. This shows that most proteins regain access within the first 15 minutes after the passage of the fork. In contrast, 25% of the identified proteins do not, and this delay cannot be inferred from their known function, physicochemical properties, or nuclear abundance. Here, we provide data where we impaired the H3K27me3 restoration to analyse its impact into chromatin restoration. To this aim, we used the EPZ-6438 inhibitor, which blocks the histone methyltransferase EZH2. Two time-points are analysed by NCC along the cell cycle in late replicated regions: nascent chromatin, just after the passage of the fork, and a sample 16 hours later, in late G1 phase. Our data support that H3K27me3 have an important role in chromatin restoration. Data available in this entry are related with the supplementary table 6 from our manuscript.

2023-02-02 | PXD038376 | Pride

RNA polymerase II acts to promote the organization of chromatin following DNA replication

Project description:In mammals, following the passage of the replisome, nucleosomes lose their defined positioning and transcription contributes to their re-organisation. However, whether transcription plays a greater role in the organization of chromatin following DNA replication remains unclear. The data uploaded are associated to the iPOND-TMT data uploaded under PXD046514 where protein re-association to newly replicated DNA was monitored upon transcription inhibition. To see whether changes observed in the iPOND-TMT time course experiments (PXD046514) are due to changes in overall protein abundance, whole extract samples were taken for each time point (Nascent, 1h and 2h after EdU labelling) and analysed by TMT-Mass Spec.

2024-02-14 | PXD040888 | Pride

Functional analysis of the replication fork proteome identifies BET proteins as PCNA regulators

Project description:Identifying proteins that function at replication forks is essential to understand DNA replication, chromatin assembly, and replication-coupled DNA repair mechanisms. Combining quantitative mass spectrometry in multiple cell types with stringent statistical cutoffs, we generated a high confidence catalogue of 593 proteins that are enriched at replication forks and nascent chromatin. Loss of function genetic analyses indicate that 85% yield phenotypes consistent with activities in DNA and chromatin replication or already have described functions in these processes. We illustrate the value of this resource by identifying activities of the BET family proteins BRD2, BRD3, and BRD4 in controlling DNA replication. These proteins use their extra-terminal domains to bind and inhibit the ATAD5 complex and thereby control the amount of PCNA on chromatin.

2020-08-17 | PXD020914 | Pride

RNA polymerase II promotes the organization of chromatin following DNA replication

Project description:Understanding how chromatin organisation is duplicated on the two daughter strands is a central question in epigenetics. In mammals, following the passage of the replisome, nucleosomes lose their defined positioning and transcription contributes to their re-organisation. However, whether transcription plays a greater role in the organization of chromatin following DNA replication remains unclear. Here we have monitored protein re-association to newly replicated DNA upon inhibition of transcription using iPOND coupled to quantitative mass spectrometry. We show that RNAPII acts to promote the re-association of hundreds of proteins with newly replicated chromatin, including ATP-dependent remodellers, transcription factors, DNA repair factors, and histone methyltransferases.

2024-02-14 | PXD046514 | Pride

RNA polymerase II acts to promote the organization of chromatin following DNA replication

Project description:In mammals, following the passage of the replisome, nucleosomes lose their defined positioning and transcription contributes to their re-organisation. However, whether transcription plays a greater role in the organization of chromatin following DNA replication remains unclear. By performing iPOND-TMT time course experiments in the presents of transcription inhibition, we show that RNAPII promotes the re-association of hundreds of proteins with newly replicated DNA (Data uploaded under PXD046514). To show that this function of RNAPII is specific to newly replicated chromatin, an iPOND-TMT 24hr chase experiment was performed in the presence of transcription inhibition to examine its effect on the protein binding on steady state chromatin.

2024-02-14 | PXD046546 | Pride

Identification of BER/SSBR proteins and the DNA glycosylases MPG, UNG2, MYH, NTH1, NEIL1 and NEIL3 in replisomes Batch 3

Project description:Base lesions in DNA can stall the replication machinery or induce mutations if bypassed. Consequently, lesions must be repaired before replication or in a post-replicative process to maintain genomic stability. Base excision repair (BER) is the main pathway for repair of base lesions, but how BER is organized during DNA replication is unclear. Here we refined the iPOND (isolation of proteins on nascent DNA) technique with targeted mass-spectrometry analysis, which enabled us to detect all proteins required for BER on nascent DNA and to monitor their spatiotemporal orchestration at replication forks. We demonstrate that XRCC1 and other BER/single-strand break repair (SSBR) proteins are enriched in replisomes in unstressed cells, supporting a cellular capacity of post-replicative BER/SSBR. Importantly, the DNA glycosylases MYH, UNG2, MPG, NTH1, NEIL1 and NEIL3 were also detected on nascent DNA. Thus our findings reveal that a broad spectrum of DNA base lesions are recognized and repaired by BER in a post-replicative process.

2024-06-07 | PXD004324 | Pride

Monitoring of the spatial and temporal dynamics of BER/SSBR pathway proteins, including MYH, UNG2, MPG, NTH1 and NEIL1-3, during DNA replication, Batch 2

2017-11-23 | PXD004494 | Pride

Monitoring of the spatial and temporal dynamics of BER/SSBR pathway proteins, including MYH, UNG2, MPG, NTH1 and NEIL1-3, during DNA replication, Batch 1

2017-11-23 | PXD004288 | Pride