{"database":"Pride","file_versions":[{"headers":{"Content-Type":["application/json"]},"body":{"files":{"Tabular":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_AXIN1Ab_PTM.tsv","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_GSK3Ab_PG_Quant.tsv","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_GSK3Ab_PG_Quant.tsv","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_AXIN1Ab_PTM.tsv","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_GSK3Ab_PTM.tsv","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_AXIN1Ab_PG_Quant.tsv","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_AXIN1Ab_PG_Quant.tsv","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_GSK3Ab_PTM.tsv"],"Raw":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_Tra_4h_AXIN1Ab_R2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_DMSO_30m_AXIN1Ab_R1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_DMSO_30m_RbIgG_R2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_DMSO_4h_RbIgG_R2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_Lysate_R2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_DMSO_30m_RbIgG_R3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_DMSO_4h_Lysate_R2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_DMSO_4h_AXIN1Ab_R3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_Chir_30m_AXIN1Ab_R4.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_DMSO_30m_RbIgG_R1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_DMSO_4h_RbIgG_R1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_DMSO_4h_Lysate_R1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_Lysate_R1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_DMSO_4h_AXIN1Ab_R2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_Lysate_R3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_Chir_30m_AXIN1Ab_R3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_Chir_30m_AXIN1Ab_R2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_DMSO_30m_AXIN1Ab_R2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_Tra_4h_AXIN1Ab_R1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_DMSO_30m_AXIN1Ab_R4.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_Tra_4h_AXIN1Ab_R3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_DMSO_4h_AXIN1Ab_R1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_Chir_30m_AXIN1Ab_R1.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_DMSO_4h_RbIgG_R3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_Lysate_R4.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/MEKi_DMSO_4h_Lysate_R3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_DMSO_30m_AXIN1Ab_R3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/06/PXD067308/GSKi_DMSO_30m_RbIgG_R4.raw"]},"type":"primary"},"statusCodeValue":200,"statusCode":"OK"}],"scores":null,"additional":{"labhead_mail":["j.krijgsveld@dkfz-heidelberg.de"],"submitter":["Robert Ihnatko"],"technology_type":["Mass Spectrometry","Data-independent acquisition"],"disease":["Colon Cancer"],"software":[""],"submitter_keywords":["Gsk3b inhibition","Immunoprecipitation","Colorectal cancer","Mek inhibition","Hct116","Lc-ms/ms"],"full_dataset_link":["https://www.ebi.ac.uk/pride/archive/projects/PXD067308"],"tissue":["Permanent Cell Line Cell","Cell Culture"],"sample_protocol":["Cells, Immunoprecipitation and Mass Spectrometry HCT116 cells were seeded on 6-well plates at a density of 20.000 cells/cm². Next day after seeding the cells were treated with 100 nM trametinib (MEK inhibitor) for 4 hours in triplicates. For GSK3B inhibition the cells were treated with 10 µM CHIR99021 (GSK3B inhibitor) for 30 min. The corresponding cells were treated with DMSO as control. Cells were then washed twice with ice-cold PBS and harvested on wet-ice using a cell scraper with 200 μl of ice-cold PBS containing protease inhibitors. The cell suspension was then pelleted by centrifugation at 1000 rpm at 4 °C. The supernatant was discarded and the cell pellets were lysed in 100 μl of RIPA buffer (Cat# 20-188 Millipore) supplemented with protease and phosphatase inhibitors (cOmplete and PhosStop, Roche). Pull-downs of either AXIN1 or GSK3B were done using Protein G Mag Sepharose (Cat# GE28-9513-79, Cytiva, Merck) conjugated with either AXIN1 or GSK3B antibody respectively, prepared in advance and kept at 4°C. For one immunoprecipitation (IP) reaction, 20 µl of Protein G Mag Sepharose conjugated with the corresponding antibody was added to the cell lysate and incubated overnight at 4 °C. Next day, the tubes with the beads were briefly spinned down, placed on a magnetic rack and incubated 5 min. The supernatant was discarded and the beads were washed 3-times with 1 mL ice-cold PBS buffer, pH 7.4 (Cat# 11503387 Fisher Scientific). After the 3rd wash, the immunoprecipitated protein complexes were stripped from the beads in two steps. In the first step, 25 µl of buffer containing 50 mM Tris pH 7.5; 1 M urea; 1 mM tris-(2-carboxyethyl)phosphine and 5 µg/mL of sequencing grade Trypsin (Promega) was added to the tubes with the beads and incubated 30 min at 27 °C on a thermomixer at 900 rpm. The tubes were then placed on a magnetic rack and incubated for another 5 minutes at room temperature (RT). The supernatant was collected into fresh, labelled tubes. In the second step, the beads were washed twice with 25 µl of buffer containing 50 mM Tris pH 7.5; 1 M urea and 5 mM chloroacetamide and the supernatant was pooled with the supernatant from the step 1 in the corresponding tubes. The tubes were incubated overnight at 27 °C to continue the digestion. Next day, the peptides generated by tryptic digestion were cleaned-up using SP3 method, as described elsewhere (Hughes et al. Nat Protocols 2019).  Preparation of antibody-conjugated Mag Sepharose beads The appropriate volume of Protein G Mag Sepharose beads calculated for the number of IP reactions was incubated with either Axin1 (C76H11) Rabbit monoclonal antibody (Cat# 2087, Cell Signaling Technology) or purified mouse Anti-GSK-3β antibody (Cat# 610202, BD Transduction Laboratories). The concentration of antibody used for conjugation was 100 µg of antibody per 100 µL of settled beads. For the corresponding IP controls, the beads were conjugated with the isotypic IgG corresponding to the Axin1 and GSK-3β antibody, in particular with either rabbit IgG (Cat# 2729, Cell Signaling Technology) or mouse (G3A1) IgG1 isotype control (Cat# 5415S, Cell Signaling Technology). The IgG to bead ratio was identical, as described above.  Liquid chromatography Tandem Mass spectrometry (LC-MS/MS)  The quantitative MS measurements were carried out using Dionex UltiMate 3000 UHPLC system (Thermo Fisher Scientific) coupled to an Exploris 480 Orbitrap mass spectrometer (Thermo Fisher Scientific). The peptides were separated by reverse-phase liquid chromatography with 0.1% formic acid (solvent A) and 100% acetonitrile supplemented with 0.1% formic acid (solvent B) as mobile phases, using a stepped gradient from 4% to 80% solvent B in 60 min on a nanoEasy M/Z peptide BEH C18 column (Waters, 250 mm x 75 μm 1/PK, 130 Å, 1.7 μm) mounted in the integrated column compartment of the UltiMate 3000 system heated to 55 °C. The peptides were eluted with a constant flow of 300 nl/min. The Exploris 480 Orbitrap mass spectrometer was operated in DIA mode with a scan range of 350-1400 m/z, orbitrap resolution 120000, normalized AGC target 300%, maxIT set to Auto mode and the precursors were analyzed in a sequence of 19 windows of variable width. The normalized HCD collision energy for the fragmentation of precursor ions was set to 28."],"repository":["Pride"],"quantification_method":[""],"modification":[""],"data_protocol":["RAW files containing spectral data were analyzed using Spectronaut™ (version 19) software using a directDIA™ workflow against a nonredundant UniProt Human Proteome FASTA database from 30.01.2020 with the identification settings as follows: precursor Q-value cutoff 0.01, precursor posterior error probability (PEP) cutoff 0.2, protein Q-value cutoff (experiment-wise) 0.01, protein Q-value cutoff (run-wise) 0.05, protein PEP cutoff 0.75. For the quantification, the data were normalized based on a retention time-dependent local regression model as described in Callister et al. J Proteome Res 2006, with precursor filtering based on identified Q-value, and maxLFQ quantification method based on interrun peptide ratios. The proteins were grouped by protein group ID and peptides were grouped by a stripped peptide sequence of the identified precursors. The missing values were imputed with 0.001 in the resulting datasets. Missing values were imputed with a low constant (0.001) to enable visualization and comparison across conditions."],"omics_type":["Proteomics"],"labhead":["Dr. Jeroen Krijgsveld"],"instrument_platform":[""],"submission_type":["PARTIAL"],"labhead_affiliation":["German Cancer Research Center (DKFZ) Heidelberg University Medical Faculty   Im Neuenheimer Feld 581 69120 Heidelberg Germany"],"species":["Homo Sapiens (human)"],"publication":["10.1186/s12964-026-02963-4","10.1186/S12964-026-02963-4"],"submitter_mail":["robert.ihnatko@gmail.com"],"submitter_affiliation":["German Cancer Research Center (DKFZ), Im Neuenheimer Feld 581, 69120 Heidelberg"],"submitter_country":["Germany"],"additional_accession":[]},"is_claimable":false,"name":"Ras-MAPK inhibition induces AXIN1 loss in colorectal cancer by mTOR associated suppression of protein synthesis.","description":"In this study, we deciphered cellular mechanisms underlying AXIN1 loss by MEK1/2 inhibition in colorectal cancer (CRC). We observed that both MEK1/2 and GSK3B inhibitors strongly reduce AXIN1 levels across multiple CRC models. Inhibition of MEK1/2, but not GSK3B, causes a reduction of AXIN1 transcript levels. While GSK3B inhibition results in rapid protein degradation of AXIN1, targeting of MEK1/2 alters neither protein stability nor post-translational modifications of AXIN1. Instead, MEK1/2 inhibition represses global translation via an mTOR dependent mechanism, which is sufficient to cause AXIN1 loss. Concordantly, mTOR inhibitors phenocopy the effect of MEK1/2 inhibitors on AXIN1 levels. Our study demonstrates that AXIN1 protein homeostasis is critically controlled by Ras-MAPK signaling at the level of protein synthesis, and that its perturbation by MEK1/2 inhibitors result in AXIN1 loss.","dates":{"publication":"2026-06-03","submission":"2025-08-13"},"accession":"PXD067308","cross_references":{"TAXONOMY":["NEWT:330879","NEWT:377960","NEWT:1131","NEWT:2042546","NEWT:259447","NEWT:295546","NCBITaxon:2719036","NEWT:1334565","NEWT:625351","NEWT:258594","NCBITaxon:1280","NEWT:112503","NEWT:1129","NEWT:309807","NEWT:59753","NEWT:89184","NEWT:309800","NEWT:281395","NEWT:1211601","NEWT:876138","NEWT:44271","NEWT:193516","NEWT:1073882","NEWT:6819","NEWT:112509","NEWT:475174","NEWT:111205","NEWT:428386","NEWT:1117","NEWT:498257","NEWT:10036","NEWT:32264","NEWT:1590","NEWT:10034","NEWT:33114","NEWT:1596","NEWT:661410","NEWT:638632","NEWT:224326","NEWT:376619","NCBITaxon:79857","NEWT:1096976","NEWT:3765","NEWT:3767","NEWT:1589","NEWT:135622","NEWT:672127","NEWT:35786","NEWT:1580","NEWT:399784","NEWT:96731","NEWT:656064","NEWT:1582","NEWT:1584","NEWT:36630","NEWT:1585","NEWT:383379","NEWT:418106","NCBITaxon:12721","NEWT:10029","NEWT:913645","NEWT:2492962","NEWT:641809","NEWT:1392488","NEWT:38815","NEWT:317447","NEWT:4688","NEWT:111225","NEWT:7719","NEWT:5530","NEWT:868565","NEWT:135674","NEWT:79329","NEWT:30069","NEWT:12637","NEWT:59729","NEWT:2164133","NEWT:295105","NEWT:108061","NEWT:60711","NEWT:224308","NEWT:3347","NEWT:49928","NEWT:160621","NEWT:212790","NEWT:110368","NEWT:55350","NEWT:1310161","NEWT:77133","NEWT:145481","NEWT:1310165","NEWT:29058","NCBITaxon:79824","NEWT:1912919","NEWT:410658","NEWT:5516","NEWT:44688","NEWT:44689","NEWT:498211","NEWT:347256","NEWT:5518","NEWT:398007","NEWT:1527468","NEWT:498217","NEWT:498216","NEWT:11320","NEWT:246196","NEWT:246197","NEWT:145458","NEWT:1310173","NEWT:44685","NEWT:161934","NEWT:156587","NEWT:1148","NEWT:5508","NEWT:3329","NEWT:419481","NEWT:5507","NEWT:410661","NEWT:1140","NCBITaxon:2157","NEWT:2066563","NEWT:1143","NEWT:1287689","NEWT:1094343","NEWT:1462472","NEWT:1336795","NEWT:644042","NEWT:150808","NEWT:1182590","NEWT:3712","NEWT:3711","NEWT:3714","NCBITaxon:37162","NEWT:270643","NEWT:2065263","NEWT:177437","NEWT:10891","NEWT:10418","NEWT:118698","NEWT:1616117","NEWT:118696","NEWT:34865","NEWT:52283","NEWT:28174","NEWT:284812","NEWT:8175","NEWT:43330","NEWT:1603293","NEWT:44664","NEWT:43346","NEWT:3702","NEWT:1245466","NEWT:3704","NEWT:244366","NEWT:1246791","NEWT:176597","NEWT:118694","NEWT:2850","NEWT:118691","NEWT:34871","NEWT:33548","NEWT:52299","NEWT:8165","NEWT:96794","NEWT:3708","NEWT:332648","NEWT:44670","NEWT:926571","NEWT:536231","NEWT:376219","NEWT:219813","NEWT:1510","NEWT:460519","NEWT:1515","NEWT:572307","NEWT:1432138","NEWT:1424507","NEWT:1194599","NEWT:272844","NEWT:1348799","NEWT:33995","NEWT:55784","NEWT:1303443","NEWT:9483","NEWT:485","NEWT:56636","NEWT:2709072","NEWT:2499525","NEWT:2853422","NEWT:487","NEWT:1538553","NEWT:1679718","NEWT:480","NEWT:67767","NEWT:46835","NEWT:109757","NEWT:582580","NEWT:1185650","NEWT:216778","NEWT:294607","NEWT:1502","NEWT:128017","NEWT:376686","NEWT:237610","NEWT:95486","EFO:0001352","NEWT:119562","NEWT:1883446","NEWT:1233435","NEWT:118223","NEWT:11757","NEWT:987059","NEWT:109760","NEWT:29031","NEWT:317047","NEWT:1509","NEWT:491","NEWT:460523","NEWT:45954","NEWT:235443","NEWT:1579","NEWT:1757312","NEWT:85694","NEWT:109786","NEWT:108458","NEWT:5936","NEWT:320637","NEWT:1078283","NEWT:3750","NEWT:983964","NEWT:11706","NEWT:32644","NEWT:527796","NEWT:499175","NEWT:109779","NEWT:3745","NEWT:1715989","NCBITaxon:4751","NEWT:3747","NEWT:1116234","NEWT:1255228","NEWT:410289","NEWT:191701","NEWT:373153","NEWT:1640278","NEWT:472","NEWT:940825","NEWT:29459","NEWT:529507","NEWT:1071661","NEWT:28128","NEWT:470","NEWT:5911","NEWT:748693","NCBITaxon:50557","NEWT:39251","NEWT:29491","NEWT:101841","NEWT:446","NEWT:153481","NEWT:2014887","NEWT:33952","NEWT:11723","NEWT:445","NEWT:153009","NEWT:261756","NEWT:166313","NEWT:63366","NEWT:63367","NEWT:215402","NEWT:1547","NEWT:1290454","NEWT:906914","NEWT:28141","NEWT:9031","NEWT:27292","NEWT:1050722","NEWT:3728","NEWT:108931","NEWT:1293497","NEWT:1055524","NEWT:548","NEWT:150475","NEWT:267872","NEWT:381666","NEWT:172269","NEWT:9534","NEWT:2126348","NEWT:5180","NEWT:256737","NEWT:9541","NEWT:8694","NEWT:33936","NEWT:8692","NEWT:2903","NEWT:185579","NEWT:13076","NEWT:207340","NEWT:1006581","NEWT:8673","NEWT:33940","NEWT:550","NEWT:33941","NEWT:554","NEWT:451516","NEWT:552","NEWT:1325291","NEWT:42410","NEWT:36185","NEWT:1054211","NEWT:1225786","NEWT:771870","NEWT:51338","NEWT:9995","NEWT:575412","NEWT:28112","NEWT:49012","NEWT:6493","NEWT:10832","NEWT:6494","NEWT:6491","NEWT:507601","NEWT:520","NEWT:186441","NEWT:643680","NEWT:214092","NCBITaxon:6157","NEWT:13095","NEWT:162425","NEWT:104105","NEWT:216257","NEWT:9986","NEWT:8654","NEWT:8658","NEWT:1268063","NEWT:8657","NEWT:9983","NEWT:8655","NEWT:5147","NEWT:595495","NEWT:28108","NEWT:28104","NEWT:407821","NEWT:568703","NCBITaxon:2","NEWT:568708","NEWT:1613","NEWT:986","NEWT:52641","NEWT:28532","NEWT:353152","NEWT:298176","NEWT:353153","NEWT:227377","NEWT:1774284","NEWT:40674","NEWT:1194669","NEWT:51329","NEWT: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