Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Identification of Plk1-PBD interacting proteins

ABSTRACT: This project aims to identify Plk1-PBD interactors potentially involved in DNA repair. We performed unbiased proteomic screens in S and G2 phases. HeLa cells were synchronized by double-thymidine block-and-release, and pull-downs were conducted with GST-Plk1 PBD as bait. GST alone and a phospho-binding-deficient GST-PBD mutant (H538A/K540M) served as negative controls. Proteins were then analyzed by mass spectrometry.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Hela Cell

DISEASE(S): Cervix Carcinoma

SUBMITTER: Shinan Zhou

LAB HEAD: Fangwei Wang

PROVIDER: PXD078902 | Pride | 2026-08-01

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	checksum.txt	Txt
	original_raw_data_GST_PBD_pulldown.raw	Raw
	original_raw_data_GST_PBDmut_pulldown.raw	Raw
	original_raw_data_GST_pulldown.raw	Raw
	search_results_data_GST_PBD_pulldown.xlsx	Xlsx

Items per page:

1 - 5 of 7

Similar Datasets

The Eyes Absent family members EYA4 and EYA1 promote PLK1 activation and successful mitosis through tyrosine dephosphorylation: siEYA4 pY IP-MS

Project description:The Eyes Absent family of proteins (EYA1-4) are a biochemically unique group of tyrosine phosphatases known to be tumour promoting across a range of cancer types. To date, the molecular targets of EYA phosphatase activity remain largely uncharacterised. Here, we identify Polo-like kinase 1 (PLK1) as a direct interactor and phosphatase substrate of both EYA4 and EYA1, with pY445 on PLK1 being the primary target site. EYA-mediated dephosphorylation of PLK1 in the G2 phase of the cell cycle is required for centrosome maturation, PLK1 localization to centrosomes, and polo-box domain (PBD) dependent interactions between PLK1 and the PLK1-activating proteins BORA and CEP192. Molecular dynamics simulations support the rationale that pY445 confers a structural impairment to PBD-substrate interactions that is relieved by EYA-mediated dephosphorylation. Depletion of EYA4 or EYA1, or chemical inhibition of EYA phosphatase activity, dramatically reduces PLK1 activation, causing mitotic defects and cell death. Overall, we have characterized a novel phosphotyrosine signalling network governing PLK1 and mitosis. This work provides a mechanism of cell killing for EYA phosphatase inhibitors with important therapeutic implications.

2024-02-26 | PXD036430 | Pride

Self-priming of Plk1 binding to BubR1 ensures accurate mitotic progression

Project description:Plk1 is a key mitotic kinase that localizes to distinct subcellular structures to promote accurate mitotic progression. Plk1 recruitment depends on direct interaction between polo-box domain (PBD) on Plk1 and PBD binding motif (PBD BM) on the interactors. However, recent study showed that PBD BM alone is not enough for stable binding between CENP-U and Plk1 highlighting the complexity of the interaction which warrants further investigation. An important interactor for Plk1 during mitosis is the checkpoint protein BubR1. Plk1 bound to BubR1 via PBD interaction with pT620 phosphorylates BubR1 S676/T680 to promote BubR1-PP2A/B56 interaction. The BubR1-PP2A/B56 complex counteracts the destablizing effect on kinetochore-microtubule attachments by mitotic kinases to promote mitotic progression. Here we show that Plk1 phosphorylates T600/T608 on BubR1 and the double phosphorylation is critical for BubR1-Plk1 interaction. A similar mechanism for Plk1-Bub1 interaction also exists indicating a general principle for Plk1 kinetochore recruitment through self-priming. Mechanistically preventing BubR1 T600/T608 phosphorylation impairs chromosome congression and checkpoint silencing by reducing Plk1 and PP2A/B56 binding to BubR1. Increasing the binding affinity towards Plk1 and PP2A/B56 in BubR1 through protein engineering bypasses the requirement of T600/T608 phosphorylation for mitotic progression. These results reveal a new layer of regulation for accurate mitotic progression.

2024-10-31 | MSV000096264 | MassIVE

Expression profiling of HeLa cells at S and G2/M phases of mitotic cell cycle.

2010-04-01 | GSE15547 | GEO

Targeting PLK1 with Onvansertib and Radiation in Head and Neck Cancer

Project description:Radiotherapy is a key treatment for head and neck squamous cell carcinoma (HNSCC) especially in patients with human papilloma virus-negative (HPV-) HNSCC. Our studies identify polo-like kinase 1 (PLK1) as a promising target for HNSCC. PLK1 inhibition with onvansertib reduces cell viability and spheroid growth of HPV- HNSCC cells. PLK1 inhibition combined with radiation increases G2/M arrest, decreases colony formation of HPV- HNSCC cells, and reduces tumor growth in vivo. RNA-sequencing demonstrates radiation increases MMP10 expression but PLK1 inhibition reverses this effect in HPV- HNSCC cells. PLK1 inhibition also reduces MMP10 activity in HPV- HNSCC cells. These findings suggest that combining PLK1 inhibition with radiation may improve therapeutic response for HPV- HNSCC via MMP10, offering a novel approach to overcome radiation resistance.

2025-10-15 | GSE306710 | GEO

Hi-C during mitotic release and G1 entry

Project description:DpnII Hi-C libraries for samples synchronized to G2/M using single 24h thymidine, 3h release, and 12h nocodazole--these samples are then released for x amount of time

2019-09-30 | GSE133462 | GEO

Expression profiling of HeLa cells at S and G2/M phases of mitotic cell cycle.

Project description:We compared the mRNAs expression profile of HeLa cells between two phases of the mitotic cell cycle: S and G2/M phases. Results provide insight into the regulation of transcript levels during mitotic cell cycle progression. HeLa cells were synchronized with double thymidine blockade (12 hours with 2 mM thymidine, 12 hours release, and 12 hours with 2 mM thymidine), and cells were taken after 2 hours release (S phase) and 8 hours release (G2/M phase). Keywords: time course Comparison of mRNAs measured at S phase (2 hours release after double thymidine blockade) and at G2/M phase (8 hours release after double thymidine blockade); 3 biological replicates at each of the two time points; two technical replicates with dye swapping per comparison; additional comparison between G2/M (8h) and S (2h).

2010-05-07 | E-GEOD-15547 | biostudies-arrayexpress

Proteomic Analysis of a PLK1-Dependent, Cell Cycle Degradation Program

Project description:Targeted protein degradation by the ubiquitin-proteasome system is an essential mechanism regulating cellular division. The kinase PLK1 coordinates protein degradation at the G2/M phase of the cell cycle by promoting the binding of substrates to the E3 ubiquitin ligase SCFβTrCP. However, the magnitude to which PLK1 shapes the mitotic proteome has not been characterized. Combining deep, quantitative proteomics with pharmacologic PLK1 inhibition (PLK1i), we identified more than 200 proteins whose abundances were increased by PLK1i at G2/M. We validate many new PLK1-regulated proteins, including several substrates of the cell cycle E3 SCFCyclin F, demonstrating that PLK1 promotes proteolysis through at least two distinct SCF-family E3 ligases. Further, we found that the protein kinase A anchoring protein AKAP2 is cell cycle regulated and that its mitotic degradation is dependent on the PLK1/βTrCP-signaling axis. Interactome analysis revealed that the strongest interactors of AKAP2 function in signaling networks regulating proliferation, including MAPK, AKT, and Hippo. Altogether, our data demonstrate that PLK1 coordinates a widespread program of protein breakdown at G2/M. We propose that dynamic proteolytic changes mediated by PLK1 integrate proliferative signals with the core cell cycle machinery during cell division. This has potential implications in malignancies where PLK1 is aberrantly regulated.

2025-05-06 | PXD046039 | Pride

Identify HILPS-interacting proteins and HIF1a phosphorylation sites

Project description:To dissect the molecular mechanisms of HILPS in promoting cell survival under hypoxia, we conducted RNA pull-down assay in combination with mass spectrometry analysis to identify HILPS-interacting proteins. Pull-down of antisense HILPS was used as a negative control. To identify the potential phosphorylation site(s), we conducted the in vitro kinase assay in combination with mass spectrometry analysis using PLK1-T210D, recombinant GST-HIF1a and cold ATP. A kinase assay using PLK1-K82R was included as a control.

2023-10-19 | PXD042226 | Pride

PLK1 phosphorylation of Epstein-Barr virus nuclear antigen 2

Project description:The Epstein-Barr virus nuclear antigen 2 (EBNA2) initiates and maintains the proliferation of infected B cells. In search of additional cellular strategies, that control EBNA2 function, we have performed a label-free mass spectrometry-based quantification of cellular proteins in EBNA2 immuno-precipitates and found polo-like kinase 1 (PLK1) to be bound to EBNA2. EBNA2/PLK1 complex formation is strongly enforced by EBNA2 S379 phosphorylation catalyzed by the mitotic CYCLIN B/CDK1 complex.

2021-08-22 | PXD022970 | Pride

Mass spectrometry analysis of in vitro PLK1 phosphorylation

Project description:Polo-like kinase 1 (PLK1) protects against genome instability by ensuring timely and accurate mitotic cell division, and its activity is tightly regulated throughout the cell cycle. Although the pathways that initially activate PLK1 in G2 are well-characterized, the factors that directly regulate PLK1 in mitosis remain poorly understood. Here, we identify that human PLK1 activity is sustained by the DNA damage response kinase Checkpoint kinase 2 (Chk2) in mitosis. Chk2 directly phosphorylates PLK1 T210, a residue on its T-loop whose phosphorylation is essential for full PLK1 kinase activity. Loss of Chk2-dependent PLK1 activity causes increased mitotic errors, including chromosome misalignment, chromosome missegregation, and cytokinetic defects. Moreover, Chk2 deficiency increases sensitivity to PLK1 inhibitors, suggesting that Chk2 status may be an informative biomarker for PLK1 inhibitor efficacy. This work demonstrates that Chk2 sustains mitotic PLK1 activity and protects genome stability through discrete functions in interphase DNA damage repair and mitotic chromosome segregation.

2024-11-01 | PXD053933 | Pride