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Hanakova"],"technology_type":["Data-dependent acquisition","Mass Spectrometry","Bottom-up proteomics"],"software":[""],"submitter_keywords":["Phosphorylation","Dishevelled","Endogenous"],"full_dataset_link":["https://www.ebi.ac.uk/pride/archive/projects/PXD063926"],"tissue":["Cell Suspension Culture"],"sample_protocol":["Samples were purified by nickel Sepharose beads, and DVL3 specific antibody in the second step of the purification and resolved on the SDS-PAGE gel. Samples were loaded onto SDS-PAGE gels, corresponding 1-D bands were excised. After destaining, the proteins in gel pieces were incubated with 10mM DTT at 56 °C for 45 min. After removal of DTT excess samples were incubated with 55mM IAA at room temperature in darkness for 30 min, then alkylation solution was removed and gel pieces were hydrated for 45 min at 4 °C in digestion solution (5 ng/µl trypsin, sequencing grade, Promega, Fitchburg, Wisconsin, USA, in 25mM AB). The trypsin digestion proceeded for 2 hours at 37 °C on Thermomixer (750 rpm; Eppendorf, Hamburg, Germany). Subsequently the tryptic digests were cleaved by chymotrypsin (5 ng/µl, sequencing grade, Roche, Basel, Switzerland, in 25mM AB) for 2 hours at 37 °C. Digested peptides were extracted from gels using 50% ACN solution with 2.5% formic acid and concentrated in speedVac concentrator (Eppendorf, Hamburg, Germany). The aliquot (1/2) of concentrated sample was transferred to LC-MS vial with already added polyethylene glycol (PEG; final concentration 0.001%) and directly analyzed by LC-MS/MS for protein identification.  The rest of sample was used for phosphopeptide analysis. Phosphopeptides were enriched using High-Select™ TiO2 Phosphopeptide Enrichment Kit (Thermo Scientific, Waltham, Massachusetts, USA) according to manufacturer protocol and extracted into LC-MS vial with already added PEG (final concentration 0.001%). Resulting peptides were analysed by LC-MS/MS. LC-MS/MS analyses of all peptide mixtures (with and without phosphoenrichment step) were done using RSLCnano system connected to Orbitrap Exploris 480 spectrometer (Thermo Fisher Scientific) with EASY Spray ion source (Thermo Fisher Scientific) installed. Prior to LC separation, tryptic digests were online concentrated and desalted using trapping column (300 μm × 5 mm, μPrecolumn, 5μm particles, Acclaim PepMap100 C18, heated compartment temperature 40°C, Thermo Fisher Scientific). After washing of trapping column with 0.1% FA, the peptides were eluted (flow 6 μl/min) from the trapping column onto Acclaim PepMap RSLC C18 column (2 µm particles, 75 μm × 250 mm; Thermo Fisher Scientific) by 74 min long gradient. Mobile phase A (0.1% FA in water) and mobile phase B (0.1% FA in 80% acetonitrile) were used in both cases. The gradient elution started at 3% of mobile phase B and increased from 3% to 37% during the first 64 min, then increased linearly to 80% of mobile phase B in the next 8 min and remained at this state for the next 2 min. Equilibration of the trapping column and the column was done prior to sample injection to sample loop.  MS data were acquired in a data-dependent strategy using survey scan (350-2000 m/z). The resolution of the survey scan was 120 000 (200 m/z) with a target value of 2.5×106 ions, one microscan and maximum injection time of 500 ms. High resolution (resolution 30 000 at 200 m/z) HCD MS/MS spectra were acquired with a target value of 2×105. Normalized collision energy was 30 % for HCD spectra. The maximum injection time for MS/MS was 250 ms. Dynamic exclusion was enabled for 45 s after one MS/MS spectra acquisition and early expiration was disabled. The isolation window for MS/MS fragmentation was set to 1.2 m/z."],"repository":["Pride"],"quantification_method":["Not available"],"modification":[""],"data_protocol":["The analysis of the mass spectrometric RAW data files was carried out using the Proteome Discoverer software (Thermo Fisher Scientific; version 1.4) with in-house Mascot (Matrixscience) search engine utilization. MS/MS ion searches were done against in-house database containing expected protein of interest with additional sequences from cRAP database (downloaded from http://www.thegpm.org/crap/). Mass tolerance for peptides and MS/MS fragments were 7 ppm and 0.03 Da, respectively. Oxidation of methionine, deamidation (N, Q), phosphorylation (S, T, Y) and carbamidomethylation of C as optional modification were set for all searches. The phosphoRS feature was used for phosphorylation localization. Peptides with false discovery rate (FDR; q-value) < 1%, rank 1 and with at least 6 amino acids were considered. Quantitative information was assessed and manually validated in Skyline software (Skyline daily)."],"omics_type":["Proteomics"],"labhead":["Zbynek Zdrahal"],"instrument_platform":[""],"labhead_affiliation":["Proteomics Core Facility and Research Group Proteomics, CEITEC-MU, Masaryk University, Brno, Czech Republic"],"submission_type":["PARTIAL"],"species":["Homo Sapiens (human)"],"submitter_mail":["katerina.hanak@gmail.com"],"publication":["Not available"],"submitter_affiliation":["CEITEC MU"],"submitter_country":["Czech Republic"],"additional_accession":[]},"is_claimable":false,"name":"Multisite phosphorylation of intrinsically disordered region of DVL","description":"When WNT binds to its receptor Frizzled (FZD) to activate the signalling, Dishevelled (DVL), the central component of Wnt signalling, is phosphorylated by redundant Casein kinase 1 (CK1) isoforms ε or δ, to transduce the signal downstream. Although the basic principles of Wnt signal transduction are known, the mechanistic aspects of DVL phosphorylation remain elusive. Here we identify Wnt-dependent multisite phosphorylation of a large and conserved serine/threonine (S/T) cluster within intrinsically disordered region (IDR), located between the PDZ and DEP domains.","dates":{"publication":"2026-04-07","submission":"2025-05-14"},"accession":"PXD063926","cross_references":{"TAXONOMY":["NEWT:6945","NEWT:184922","NEWT:6703","NEWT:3555","NEWT:2","NEWT:157546","NEWT:35554","NEWT:38942","NEWT:307972","NEWT:32046","NEWT:544496","NEWT:2102","NEWT:2042546","NEWT:45351","NEWT:43179","NEWT:4513","NEWT:5722","NEWT:1247","NEWT:55153","NCBITaxon:10407","NEWT:1736309","NEWT:309800","NEWT:281395","NEWT:10360","NEWT:1211601","NEWT:876138","NEWT:47664","NEWT:3654","NEWT:237561","NEWT:5833","NEWT:6928","NEWT:10036","NEWT:36745","NEWT:498019","NEWT:1351","NEWT:1438992","NEWT:2649997","NEWT:272563","NEWT:224326","NCBITaxon:79857","NEWT:1096976","NEWT:82688","NEWT:95648","NEWT:3885","NEWT:3888","NEWT:5821","NEWT:1589","NEWT:135622","NCBITaxon:4896","NEWT:6915","NEWT:3649","NEWT:101510","NEWT:28903","NEWT:3880","NEWT:272559","NEWT:28909","NEWT:3641","NEWT:383379","NEWT:466585","NEWT:10029","NEWT:913645","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:3347","NEWT:139927","NEWT:4558","NEWT:209285","NEWT:5888","NEWT:211586","NEWT:1283","NEWT:931281","NEWT:4550","NEWT:1000561","NEWT:197","NEWT:1390363","NEWT:288705","NCBITaxon:79824","NEWT:4787","NCBITaxon:4563","NEWT:5755","NEWT:44689","NEWT:3218","NEWT:5759","NEWT:1736231","NEWT:1270","NEWT:374990","NEWT:498217","NEWT:2242","NEWT:4784","NEWT:11320","NEWT:360106","NEWT:286","NEWT:391619","NEWT:360104","NEWT:287","NEWT:246197","NEWT:10117","NEWT:10239","NEWT:10116","NEWT:1280","NEWT:1735272","NEWT:83334","NEWT:83332","NEWT:44685","NEWT:317513","NEWT:1148","NEWT:580240","NEWT:5508","NEWT:294128","NEWT:11676","NEWT:55571","NEWT:35500","NEWT:1140","NEWT:100226","NEWT:4530","NEWT:4896","NEWT:75058","NEWT:13616","NEWT:1390","NEWT:1094343","NEWT:1336795","NEWT:296543","NEWT:1773","NEWT:1895","NEWT:1182590","NEWT:3712","NEWT:82380","NEWT:105023","NEWT:935293","NEWT:64152","NEWT:4924","NEWT:749200","NEWT:375146","NEWT:990346","NEWT:145953","NEWT:257309","NEWT:100816","NEWT:263","NEWT:230741","NEWT:52283","NEWT:284812","NCBITaxon:1313","NEWT:43330","NEWT:1603293","NEWT:408169","NEWT:44544","NEWT:47946","NEWT:4911","NEWT:645463","NEWT:3702","NEWT:129249","NEWT:243277","NEWT:990119","NEWT:408172","NEWT:408170","NEWT:493760","NEWT:106590","NEWT:260710","NEWT:257313","NEWT:400772","NEWT:3708","NEWT:128161","NEWT:332648","NEWT:106592","NEWT:536231","NEWT:1436733","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:109757","NEWT:943146","NEWT:2711","NEWT:300852","NEWT:1502","NEWT:376686","NEWT:95486","NEWT:9103","NEWT:1883446","NEWT:29159","NEWT:253","NEWT:10306","NCBITaxon:2759","NEWT:1233435","NEWT:93061","NEWT:8022","NEWT:145943","NCBITaxon:4932","NEWT:595536","NEWT:240906","NEWT:593117","NEWT:89920","NEWT:3635","NEWT:5811","NEWT:235443","NEWT:108458","NEWT:272623","NEWT:272624","NEWT:411483","NEWT:884019","NEWT:198215","NEWT:411490","NEWT:983964","NEWT:118499","NEWT:169963","NEWT:32644","NEWT:225117","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:1071661","NEWT:360094","NEWT:470","NEWT:41364","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:38293","NEWT:327159","NEWT:178876","NEWT:327160","NEWT:573","NEWT:9031","NEWT:1872122","NEWT:7091","NEWT:108931","NEWT:241368","NEWT:42528","NEWT:190802","NEWT:9778","NEWT:150475","NEWT:303","NEWT:9417","NEWT:7111","NEWT:347515","NEWT:1216979","NEWT:7237","NEWT:5180","NEWT:256737","NEWT:9541","NEWT:115104","NEWT:1121114","NEWT:663","NEWT:1081927","NEWT:1238993","NEWT:67825","NEWT:185579","NEWT:941442","NEWT:220668","NEWT:13076","NEWT:1249668","NEWT:317","NEWT:7227","NEWT:7469","NEWT:885318","NEWT:9402","NEWT:9644","NEWT:415540","NEWT:550","NEWT:675060","NEWT:4081","NEWT:334542","NEWT:554","NEWT:27592","NEWT:98334","NEWT:426428","NEWT:588858","NEWT:9639","NEWT:242231","NEWT:7574","NEWT:1715256","NEWT:7215","NEWT:575412","NEWT:929793","NEWT:29204","NEWT:2172103","NEWT:6494","NEWT:7460","NEWT:6491","NEWT:507601","NEWT:643680","NCBITaxon:6157","NEWT:42752","NEWT:746360","NEWT:6239","NEWT:470150","NEWT:162425","NEWT:216257","NEWT:102169","NEWT:543734","NEWT:9986","NEWT:4054","NEWT:73239","NEWT:226186","NEWT:1268063","NEWT:8782","NEWT:1263854","NEWT:118503","NEWT:435590","NEWT:1902","NEWT:160488","NEWT:28104","NEWT:1908","NEWT: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