{"database":"Pride","file_versions":[{"headers":{"Content-Type":["application/json"]},"body":{"files":{"Txt":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/peptides.txt","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/proteinGroups.txt","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/checksum.txt"],"Xml":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/mqpar.xml"],"Raw":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F3_Split_1_092513.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F5_Split_2_093555.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F3_Split_2_092808.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F2_Split_2_092407.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F5_Split_1_093313.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F6_Split_1_093719.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F2_Split_1_092051.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F4_Split_2_093208.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F4_Split_1_092908.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F1_Split_2_091956.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F1_Split_1_091653.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD061002/TMT_NCDN_F6_Split_2_093948.raw"]},"type":"primary"},"statusCode":"OK","statusCodeValue":200}],"scores":null,"additional":{"labhead_mail":["r.barderasm@isciii.es"],"submitter":["Ana Montero Calle"],"technology_type":["Data-dependent acquisition","Mass Spectrometry","Bottom-up proteomics"],"disease":["Colon Cancer"],"software":[""],"submitter_keywords":[""],"full_dataset_link":["https://www.ebi.ac.uk/pride/archive/projects/PXD061002"],"tissue":["Epithelial Cell"],"sample_protocol":["Cells at confluence and maintained for 48 h in serum-free media were harvested with 4mM EDTA-PBS and centrifuged at 260 g and RT for 5 min. Then, cell pellets were lysed with 500 µL of RIPA buffer supplemented with protease and phosphatase inhibitors.  For the TMT analyses, 10-plex TMT analyses of NCDN-depleted cells were performed with KM12C and KM12SM cells. Both control and silenced KM12C and KM12SM cells were labeled in duplicate.  Briefly, a total of 10 µg of each protein extract or pool in 100 µl of RIPA were reduced with 10 µl 100 mM TCEP for 45 minutes at 37°C and 600 rpm and alkylated with 11 µL of 400 mM chloroacetamide for 30 minutes at RT, 600 rpm and in darkness. After that, samples were incubated with 100 µl of SeraMag magnetic beads mix (50% hydrophilic beads - 50% hydrophobic beads) and 200 µl of 100 % acetonitrile (ACN) for 35 minutes at RT and 600 rpm. Then, the supernatants were discarded, and the magnetic beads with the bound proteins were washed twice with 70% ethanol and once with ACN. Lastly, supernatants were discarded, proteins were O/N digested at 37°C and 600 rpm with 0.5 µg of porcine trypsin in 100 µL of 200 mM HEPES, pH 8.0. Next, samples were sonicated twice, and the supernatants containing digested peptides were collected. Samples were separately labeled in duplicate with the different Tandem Mass Tags in two incubation steps of 30 minutes at RT and 600 rpm with 10 µl of reagent per incubation, and a final incubation with 10 µl of 1 M glycine, pH 2.7 for 30 minutes at RT and 600 rpm. Finally, the content of the 10 tubes was pooled together and dried under vacuum prior to peptide separation using High pH Reversed-Phase Peptide Fractionation Kit (Thermo Fisher Scientific). For separation, dried peptides were reconstituted in 300 µl of 0.1% TFA in H2Omq and columns were equilibrated twice with 300 µl of ACN and twice with 300 µl of 0.1% TFA in H2Omq. Then, peptides were loaded to the columns, washed twice with 300 µl of 0.1% TFA in H2Omq, and separated in 12 fractions of 300 µl each in 0.1% triethylamine, 2.5-100% ACN. Then, fractions were dried under vacuum and stored at -80°C until analysis by LC-MS/MS. Samples were reconstituted in 10 µl of 0.1% FA prior to LC-MS/MS analysis. TMT experiments were analyzed on an Orbitrap Exploris 480 mass spectrometer (Thermo Fisher Scientific) equipped with FAIMS Pro Duo interface technology following laboratory’s standard guidelines. For FAIMS, a gas flow of 4.6 L/min and CVs -70V and -50V were used. For each analysis, samples were loaded into the precolumn PepMap 100 C18 3 μm, 75 μm x 2 cm Nanoviper Trap 1200 BA (Thermo Fisher Scientific), and eluted at 50 °C on an Easy-Spray PepMap RSLC C18 2 μm, 75 μm x 50 cm (Thermo Fisher Scientific). The flow rate for the mobile phase was 300 nl/min using 0.1% formic acid in water as solvent A, and 0.1% formic acid in 80% acetonitrile as solvent B. The set gradient profile was: 0%-2% buffer B for 4 min, 2% buffer B for 2 min, 2%-42% buffer B for 100 min, 42%-72% buffer B for 14 min, 72%-95% buffer B for 5 min and 95% buffer B for 10 min. For ionisation, 2300V liquid junction voltage and 280 °C of capillary temperature were set. The full scan method employed a mass selection m/z 350-1400, a resolution of 60000 (at m/z 200), an automatic gain control (AGC) of 300% and a maximum injection time of 25 ms. After survey scan, the 12 most intense precursor ions were selected for MS/MS fragmentation, which was performed with a normalized collision energy of 32, a starting mass m/z 100, a 100% AGC target, a resolution of 15000 (at m/z 200), an intensity threshold of 2x104, an isolation window of 0.7 m/z units, a maximum IT of 22 ms and the TurboTMT enabled. Charge state screening was used for reject unassigned, singly charged, and greater than or equal to 7 protonated ions. The dynamic exclusion time used to discriminate against previously selected ions was 30 s."],"repository":["Pride"],"quantification_method":[""],"modification":[""],"data_protocol":["For data analysis MaxQuant (version 2.1.3) was used using standardized workflows. Mass spectra files were searched against Uniprot UP000005640_9606.fasta Homo sapiens (human) 2019 database (20962 protein entries) using the MS2 type reporter ion. Precursor and reporter mass tolerance was set at 4.5 ppm and 0.003 Da respectively. MS data were analyzed with MaxQuant (version 1.6.6.0 and 2.1.3) using standardized workflows. TMTs data normalization and sample loading (SL) normalization was performed with R Studio (version 4.1.1) according to established protocols (https://github.com/pwilmart). For statistical analysis of TMTs, moderated t-statistics analysis was performed also with R Studio (version 4.1.1) according to laboratory’s guidelines. Proteins identified with 1 or more unique peptides, a fold change of ≥ 1.5 or ≤ 0.67 and p value < 0.05, were selected as up-regulated and down-regulated, respectively."],"omics_type":["Proteomics"],"labhead":["Rodrigo Barderas"],"instrument_platform":[""],"labhead_affiliation":["Chronic Disease Programme (UFIEC), Instituto de Salud Carlos III, Madrid, Spain."],"submission_type":["PARTIAL"],"species":["Homo Sapiens (human)"],"submitter_mail":["ana.monteroc@isciii.es"],"publication":["10.1038/S41419-026-08747-5"],"submitter_affiliation":["Instituto de Salud Carlos III"],"submitter_country":["Spain"],"additional_accession":[]},"is_claimable":false,"name":"Neurochondrin Drives Colorectal Cancer Progression by Modulating the PODXL–Ezrin Axis and Mitochondrial Function","description":"Neurochondrin (NCDN) has been recently identified as upregulated in metastatic to liver KM12SM colorectal cancer (CRC) cells in comparison to the poorly metastatic KM12C CRC cells. Furthermore, high levels of NCDN in patients correlate to a worse survival. In this work, using an independent patient cohort, we validated the worsened prognosis associated to NCDN’s elevated levels. Then, we sought to investigate the role of NCDN in CRC progression using the isogenic KM12 CRC cell model comprised of the aforementioned KM12C and KM12SM cells. Stable silencing of NCDN in both cell lines produced a reduction on the tumorigenic capacities of the cells in vitro as seen through different assays (proliferation, adhesion, colony formation…). In vivo, mice injected with the silenced cells had impaired tumor growth and metastasis compared to mice injected with the control cells. Proteomic profiling of NCDN-silenced cells uncovered a set of dysregulated proteins associated with NCDN such as PODXL, indicating that the NCDN-PODXL axis constitutes a critical mediator of CRC liver metastasis. Our findings position NCDN and their associated dysregulated proteins as drivers of metastatic progression in CRC.","dates":{"publication":"2026-05-03","submission":"2025-02-20"},"accession":"PXD061002","cross_references":{"TAXONOMY":["NEWT:1773","NEWT:3555","NEWT:1182590","NEWT:10090","NEWT:749200","NEWT:35554","NEWT:4120","NEWT:5693","NEWT:347515","NEWT:1216979","NEWT:307972","NEWT:92867","NEWT:990346","NEWT:544496","NEWT:145953","NEWT:284812","NEWT:115104","NEWT:43330","NEWT:67825","NEWT:44544","NEWT:13076","NEWT:544404","NEWT:3702","NEWT:8839","NEWT:4232","NEWT:1736309","NEWT:4113","NEWT:7227","NEWT:11298","NEWT:885318","NEWT:4081","NEWT:876138","NEWT:554","NEWT:5691","NEWT:260710","NEWT:106592","NEWT:237561","NEWT:9913","NEWT:10036","NEWT:4100","NEWT:7574","NEWT:1351","NEWT:1076","NEWT:6763","NEWT:7215","NEWT:380394","NEWT:272563","NEWT:1639","NEWT:188229","NEWT:746360","NEWT:6239","NEWT:135588","NEWT:135622","NEWT:6915","NEWT:9986","NEWT:101510","NEWT:3880","NEWT:58002","NEWT:9103","NEWT:4577","NEWT:146479","NEWT:1000589","NEWT:145943","NEWT:85962","NEWT:160488","NEWT:317447","NEWT:3635","NEWT:7955","NCBITaxon:2","NEWT:7959","NEWT:2261","NEWT:3197","NEWT:9615","NEWT:884019","NEWT:4565","NEWT:1264690","NEWT:169963","NEWT:36329","NEWT:34305","NEWT:59729","NEWT:626528","NEWT:139927","NEWT:4558","NEWT:9606","NEWT:367830","NEWT:243230","NEWT:931281","NEWT:7029","NEWT:1283300","NEWT:334747","NEWT:470","NEWT:3218","NEWT:5759","NEWT:9838","NCBITaxon:9615","NEWT:1736231","NEWT:1193501","NEWT:6287","NEWT:6326","NEWT:9796","NEWT:2762","NEWT:5476","NEWT:562","NEWT:260707","NEWT:287","NEWT:10117","NEWT:10116","NEWT:1280","NEWT:1836","NEWT:29760","NEWT:260705","NEWT:1148","NEWT:4932","NEWT:70448","NEWT:9825","NEWT:3603","NEWT:698936","NEWT:39946","NEWT:11676","NEWT:9823","NEWT:100226","NCBITaxon:6073","NEWT:4896","NEWT:6279","NEWT:7370","NEWT:573","NEWT:6282","NEWT:7091"],"pubmed":[],"ORCID":["0000-0001-5141-0454"]}}