{"database":"Pride","file_versions":[{"headers":{"Content-Type":["application/json"]},"body":{"files":{"Raw":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/1B-p_20191016135704.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/1C-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/4B-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/6B-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/5B-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/2C-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/4C-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/3C-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/1A-p_20191016123902.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/3D-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/2D-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/5C-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/6C-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/5D-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/4D-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/1D-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/6A-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/2A-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/2B-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/3B-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/3A-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/5A-p.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/4A-p.raw"],"Other":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/10/PXD065997/combined.rar"]},"type":"primary"},"statusCode":"OK","statusCodeValue":200}],"scores":null,"additional":{"labhead_mail":["libor.macurek@img.cas.cz"],"submitter":["Pavel Talacko"],"technology_type":["Mass Spectrometry","Bottom-up proteomics"],"software":[""],"submitter_keywords":["Rna polymerase i","Phosphorylation","Tcof1","Nucleolus","Condensation"],"full_dataset_link":["https://www.ebi.ac.uk/pride/archive/projects/PXD065997"],"tissue":["Cell Culture"],"sample_protocol":["Cells were lysed by boiling at 95°C for 10 min in 100mM Tris pH 8.5 containing 2% SDC (sodium deoxycholate), 40mM  Chloroacetamide and 10 mM TCEP (Tris(2-carboxyethyl)phosphine) and further sonicated (Bandelin Sonoplus Mini 20, MS 1.5). Protein concentration was determined using BCA protein assay kit (Thermo) and 250 µg of protein per sample was used for MS sample preparation. Proteins were digested by 5 µg of trypsin per sample at 37°C overnight. Phosphopeptides were enriched using TiO2 according to Humphrey et al.Nano Reversed phase columns (EASY-Spray column, 50 cm x 75 µm ID, PepMap C18, 2 µm particles, 100 Å pore size) were used for LC/MS analysis. Mobile phase buffer A was composed of water and 0.1% formic acid. Mobile phase B was composed of acetonitrile and  0.1% formic acid. Samples were loaded onto the trap column (C18 PepMap100, 5 μm particle size, 300 μm x 5 mm, Thermo Scientific) for 4 min at 18 μl/min loading buffer was composed of water, 2% acetonitrile and 0.1% trifluoroacetic acid. Peptides were eluted with Mobile phase B gradient from 2% to 35% B in 60 min. Eluting peptide cations were converted to gas-phase ions by electrospray ionization and analyzed on a Thermo Orbitrap Fusion (Q-OT- qIT, Thermo Scientific). Survey scans of peptide precursors from 350 to 1400 m/z were performed in orbitrap at 120K resolution (at 200 m/z) with a 1 × 106 ion count target. Tandem MS was performed by isolation at 1,5 Th with the quadrupole, HCD fragmentation with normalized collision energy of 35, and rapid scan MS analysis in the ion trap. The MS2 ion count target was set to 104 and the max injection time was 150 ms. Only those precursors with charge state 2–6 were sampled for MS2. The dynamic exclusion duration was set to 30 s with a 10ppm tolerance around the selected precursor and its isotopes. Monoisotopic precursor selection was turned on. Cycle time was se to 2 s."],"repository":["Pride"],"quantification_method":["Not available"],"modification":[""],"data_protocol":["All data were analyzed and quantified with the MaxQuant software (version 2.0.1.0) (Cox, Mann 2008). The false discovery rate (FDR) was set to 1% for both proteins and peptides. The Andromeda search engine was used for the MS/MS spectra search against the Human database (downloaded from Uniprot). Enzyme specificity was set as C-terminal to Arg and Lys, also allowing cleavage at proline bonds and a maximum of two missed cleavages. Carbamidomethylation of cysteine was selected as fixed modification and N- terminal protein acetylation and methionine oxidation as variable modifications. The “match between runs” feature of MaxQuant was used to transfer identifications to other LC-MS/MS runs based on their masses and retention time (maximum deviation 0.7 min) and this was also used in quantification experiments. Quantifications were performed with the label-free algorithm in MaxQuant (Cox et al. 2014). Data analysis was performed using Perseus 1.6.15.0 software (Tyanova et al.). Only those phosphosites with localization probability higher than 0.75 were used for further data analysis."],"omics_type":["Proteomics"],"labhead":["Libor Macurek"],"instrument_platform":[""],"submission_type":["PARTIAL"],"labhead_affiliation":["Cancer Cell Biology, Institute of Molecular Genetics of the Czech Academy of Sciences, Prague, CZ-14220, Czech Republic"],"species":["Homo Sapiens (human)"],"publication":["Not available"],"submitter_mail":["pavel.talacko@natur.cuni.cz"],"submitter_affiliation":["BIOCEV, Faculty of Science, Charles University in Prague"],"submitter_country":["Czechia"],"additional_accession":[]},"is_claimable":false,"name":"Dynamic phosphorylation of TCOF1 controlled by CK2 and PPM1D regulates nucleolar condensation and RNA Polymerase I activity","description":"The nucleolus is a membrane-less organelle essential for rRNA transcription and ribosome biogenesis, yet the mechanisms regulating its dynamic assembly remain incompletely understood. Here, we identify the phosphatase PPM1D as a key regulator of the nucleolar architecture via dephosphorylation of the scaffold protein TCOF1. PPM1D localizes to the nucleolus through a conserved motif at the C-terminus and targets CK2-phosphorylated sites within the low-complexity repeat domain of TCOF1. These phosphorylation events promote TCOF1 condensation, immobilization in the fibrillar centres, and interaction with the RNA polymerase I, supporting rRNA transcription. Disruption of the phosphorylation balance by loss of PPM1D or inhibition of CK2 impairs TCOF1 function, leading to nucleolar disorganization and reduced rRNA transcription. Structural modelling supports a phosphorylation-dependent interaction between TCOF1 and POLR1A. Furthermore, we demonstrate that pharmacological inhibition of PPM1D sensitizes cells to RNA polymerase I inhibitors, in both p53-dependent and independent contexts. Together, these findings uncover a reversible phosphorylation switch that governs TCOF1 dynamics and its nucleolar function, with potential implications for targeting ribosome biogenesis in cancer.","dates":{"publication":"2026-10-02","submission":"2025-07-10"},"accession":"PXD065997","cross_references":{"TAXONOMY":["NEWT:6945","NEWT:3555","NEWT:2","NEWT:157546","NEWT:35554","NEWT:38942","NEWT:307972","NEWT:32046","NEWT:544496","NEWT:2042546","NEWT:45351","NEWT:43179","NEWT:4513","NEWT:5722","NEWT:376741","NEWT:55153","NCBITaxon:10407","NEWT:1736309","NEWT:309800","NEWT:1211601","NEWT:876138","NEWT:557436","NEWT:237561","NEWT:5833","NEWT:6928","NEWT:10036","NEWT:36745","NEWT:1351","NEWT:1438992","NEWT:2649997","NEWT:272563","NEWT:79220","NEWT:224326","NCBITaxon:79857","NEWT:1096976","NEWT:95648","NEWT:3885","NEWT:3888","NEWT:1589","NEWT:135622","NCBITaxon:4896","NEWT:6915","NEWT:3649","NEWT:101510","NEWT:3880","NEWT:272559","NEWT:3641","NEWT:383379","NEWT:466585","NEWT:10029","NEWT:1000589","NEWT:85963","NEWT:85962","NEWT:317447","NEWT:7955","NEWT:7959","NEWT:2261","NEWT:31156","NEWT:398580","NEWT:4565","NEWT:1264690","NEWT:515619","NEWT:192875","NEWT:34305","NEWT:59729","NCBITaxon:183674","NEWT:224308","NEWT:84645","NEWT:626528","NEWT:139927","NEWT:4558","NEWT:209285","NEWT:211586","NEWT:1283","NEWT:931281","NEWT:4550","NEWT:1000561","NEWT:197","NCBITaxon:79824","NEWT:4787","NCBITaxon:4563","NEWT:5755","NEWT:44689","NEWT:3218","NEWT:5759","NEWT:1736231","NEWT:1270","NEWT:2242","NEWT:4784","NEWT:11320","NEWT:360106","NEWT:286","NEWT:287","NEWT:10117","NEWT:10239","NEWT:10116","NEWT:1280","NEWT:1735272","NEWT:83334","NEWT:83332","NEWT:44685","NEWT:317513","NEWT:1148","NEWT:580240","NEWT:294128","NEWT:11676","NEWT:55571","NEWT:100226","NEWT:4530","NEWT:4896","NEWT:75058","NEWT:13616","NEWT:1094343","NEWT:296543","NEWT:1773","NEWT:1895","NEWT:1182590","NEWT:3712","NEWT:935293","NEWT:64152","NEWT:4924","NEWT:749200","NEWT:990346","NEWT:145953","NEWT:257309","NEWT:100816","NEWT:263","NEWT:230741","NEWT:52283","NEWT:284812","NCBITaxon:1313","NEWT:43330","NEWT:980415","NEWT:1603293","NEWT:408169","NEWT:44544","NEWT:4911","NEWT:645463","NEWT:3702","NEWT:129249","NEWT:243277","NEWT:990119","NEWT:408172","NEWT:408170","NEWT:493760","NEWT:260710","NEWT:257313","NEWT:400772","NEWT:3708","NEWT:128161","NEWT:332648","NEWT:106592","NEWT:536231","NEWT:460519","NEWT:1187947","NEWT:1432138","NEWT:10312","NEWT:1424507","NCBITaxon:1773","NEWT:9598","NEWT:8030","NEWT:1639","NEWT:188229","NEWT:3818","NEWT:480","NEWT:4909","NEWT:67767","NEWT:432359","NEWT:46835","NEWT:1182263","NEWT:2711","NEWT:376686","NEWT:95486","NEWT:9103","NEWT:29159","NEWT:253","NEWT:10306","NCBITaxon:2759","NEWT:1233435","NEWT:8022","NEWT:145943","NCBITaxon:4932","NEWT:595536","NEWT:240906","NEWT:593117","NEWT:3635","NEWT:5811","NEWT:235443","NEWT:272624","NEWT:2719099","NEWT:411483","NEWT:884019","NEWT:198215","NEWT:411490","NEWT:983964","NEWT:169963","NEWT:32644","NEWT:499175","NEWT:109779","NEWT:476272","NEWT:3747","NEWT:195051","NEWT:367830","NEWT:1255228","NEWT:178616","NEWT:410289","NEWT:373153","NEWT:352472","NEWT:357","NEWT:360094","NEWT:470","NEWT:1313","NEWT:411469","NEWT:84023","NEWT:559292","NEWT:39491","NCBITaxon:5811","NEWT:411464","NEWT:411460","NEWT:2014887","NEWT:2762","NEWT:1174673","NEWT:562","NEWT:411470","NEWT:33952","NEWT:2094720","NCBITaxon:2697049","NEWT:571256","NEWT:28038","NEWT:1663","NEWT:1423","NEWT:4932","NEWT:3603","NEWT:2759","NEWT:3847","NEWT:327159","NEWT:178876","NEWT:327160","NEWT:573","NEWT:9031","NEWT:7091","NEWT:241368","NEWT:42528","NEWT:190802","NEWT:9778","NEWT:150475","NEWT:303","NEWT:9417","NEWT:7111","NEWT:347515","NEWT:1216979","NEWT:5180","NEWT:256737","NEWT:115104","NEWT:1121114","NEWT:663","NEWT:1081927","NEWT:1238993","NEWT:67825","NEWT:185579","NEWT:941442","NEWT:220668","NEWT:13076","NEWT:1249668","NEWT:7108","NEWT:317","NEWT:7227","NEWT:7469","NEWT:885318","NEWT:9402","NEWT:415540","NEWT:550","NEWT:675060","NEWT:4081","NEWT:334542","NEWT:554","NEWT:98334","NEWT:426428","NEWT:7574","NEWT:1715256","NEWT:7215","NEWT:575412","NEWT:29204","NEWT:2172103","NEWT:507601","NEWT:643680","NCBITaxon:6157","NEWT:746360","NEWT:6239","NEWT:470150","NEWT:216257","NEWT:102169","NEWT:9986","NEWT:4054","NEWT:73239","NEWT:226186","NEWT:1268063","NEWT:8782","NEWT:1263854","NEWT:435590","NEWT:1902","NEWT:160488","NEWT:28104","NEWT:1908","NCBITaxon:2","NEWT:985076","NEWT:1215323","NEWT:623","NEWT:52641","NEWT:7038","NEWT:6192","NEWT:28532","NCBITaxon:38727","NEWT:198618","NEWT:353152","NEWT:2829","NEWT:366581","NEWT:216599","NEWT:216595","NEWT:51329","NEWT:243230","NEWT:8355","NEWT:9685","NEWT:7029","NEWT:1080772","NEWT:1283300","NEWT:6183","NEWT:6063","NEWT:334747","NEWT:61235","NEWT:6289","NEWT:436486","NEWT:6287","NEWT:300641","NEWT:727","NEWT:9796","NEWT:725","NEWT:170187","NEWT:260707","NCBITaxon:6191","NEWT:1836","NEWT:185431","NEWT:29760","NEWT:260704","NEWT:703612","NEWT:260705","NEWT:80863","NEWT:2697049","NEWT:105231","NEWT:1216981","NCBITaxon:6073","NEWT:884204","NEWT:6279","NEWT:1123869","NEWT:9544","NEWT:7370","NEWT:83906","NEWT:607699","NEWT:6282","NEWT:426688","NEWT:208964","NEWT:1134506","NEWT:5755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