{"database":"Pride","file_versions":[{"headers":{"Content-Type":["application/json"]},"body":{"files":{"Mzml":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/09/PXD047995/Organoides_Spectra.mzML"],"Raw":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/09/PXD047995/20211029_9_TMT_Fr3.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/09/PXD047995/20211029_12_TMT_Fr4.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/09/PXD047995/20211029_6_TMT_Fr2.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/09/PXD047995/20211029_15_TMT_Fr5.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/09/PXD047995/20211029_3_TMT_Fr1.raw"],"Mzid":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/09/PXD047995/organoides_mz2.mzid.gz"]},"type":"primary"},"statusCode":"OK","statusCodeValue":200}],"scores":null,"additional":{"labhead_mail":["fcorrales@cnb.csic.es"],"submitter":["Irene Blázquez García"],"technology_type":["Data-dependent acquisition","Mass Spectrometry","Bottom-up proteomics"],"disease":["Progressive Familial Intrahepatic Cholestasis 3"],"software":[""],"submitter_keywords":["Mdr2","Pfic3","Organoid","Cholestasis and hepatocellular carcinoma"],"full_dataset_link":["https://www.ebi.ac.uk/pride/archive/projects/PXD047995"],"tissue":["3-d Shape"],"sample_protocol":["Aliquots of organoid lysates containing 50 ï�g protein were loaded into micro S-Trap columns (Protifi). Reduction and alkylation of Cys sulfhydryl groups was then performed by incubation with tris (2- carboxyethyl) phosphine (TCEP, 5mM) and chloroacetamide (CCA, 10 mM), for 1h at 37°C under agitation. Protein digestion was performed upon addition of trypsin (90057, ThermoFisher Scientific) at a 1:15 ratio (µg trypsin: µg protein) o/n at 37ºC12. The resulting peptide concentration was determined the fluorometric Qubit assay (Invitrogen).  Isobaric labeling of the tryptic peptides was performed with the TMT-6plex kit (TMTsixplexâ„¢ Isobaric Label Reagent Set, 90057, ThermoFisher Scientific, supplementary table 3). 35 µg peptides from each sample were resuspended in 100 µL of labeling buffer (50% acetonitrile, ACN/25 mM TEAB) along with its corresponding label tag. The labeling reaction was carried out for 2 hours at room temperature under stirring (600 rpm). The reaction was stopped by incubation at room temperature for 15 min with 0.3% hydroxylamine. Then, all labeled samples were mixed and an aliquot containing 80 µg peptides was taken for peptide pre-fractionation. The resulting aliquots were dried in Speed-vac and stored at -20°C. 80 µg of labeled peptides were resuspended in 100 µL 1% formic acid (FA). Solid phase fractionation was carried out in CDS Emporeâ„¢ SDB-RPS Extraction Disks (13-110-022, Fisher scientific) at basic pH due to the presence of ammonium formate (NH4HCO2) and at increasing concentrations of ACN. Finally, the eluted fractions were combined together (1-6, 2-7, 3-8, 4-9, 5-10) and dried in the Speed-vac.  One µg of the tryptic digest was analyzed by 1D-nano LC (Ultimate 3000 nano HPLC, ThermoFisher Scientific) coupled to Orbitrap Explorisâ„¢ 240 mass spectrometer (Thermo Fisher Scientific). Peptides were separated on an Easy-spray PepMap C18 analytical column (50 cm × 75 μm) at 45°C by applying a flow rate of 250 nL/min with a 120 min gradient from 4 % to 95 % mobile phase B (Mobile phase A: 0.1% FA; mobile phase B: 80 % ACN in 0.1% FA). In the last 10 min of this gradient, the ratio of each phase returns to the initial conditions (96% Phase A and 4% Phase B). The sample loading buffer was 2% ACN in 0.1% FA and the injection volume of the equipment was 5 µL."],"repository":["Pride"],"quantification_method":[""],"modification":[""],"data_protocol":["Data acquisition was carried out with the Data Dependent Acquisition (DDA) method, in positive mode, monitoring all ions in a range of 375 to 1200 mass/charge (m/z). The 20 most intense ions from each MS1 scan were selected and fragmented by Higher-energy collisional dissociation (HCD). The resolution of each spectrum after fragmentation was set at 45,000 at m/z 200, the precursor isolation window at 0.7 m/z, with an exclusion time of 45 s and the HCD collision energy at 30. Precursor ions that showed one or no assignment after fragmentation were excluded, as well as those with a charge of 6 or higher. Raw mass spectrometry data were processed with the Proteome Discoverer (v2.5, ThermoFisher Scientific) using four search engines (Mascot (v2.7.0), MsAmanda (v2.4.0), MsFragger (v3.1.1) and Sequest HT) and two databases (target/decoy) constructed from SwissProt Mus musculus sequences (Tax ID: 10090, 25097 sequences, 09/22/2021). Two types of searches were performed depending on the configuration of the TMT modification (+229,163 Da in lysine and N-terminal peptides): 1) Dynamic search, if this modification was set as a variable modification, which is used to calculate the efficiency of TMT tagging; 2) Static search, if it was set as a fixed modification, which is used to quantify tagged peptides. The rest of the search parameters were, fixed modification, carbamidomethylation (+57,021 Da); variable modificatioins, Gln-pyroGlu (+17,027 Da), methionine oxidation (+15,9949 Da), N-terminal acetylation (+42,011 Da); enzyme, trypsin and 2 missed cleavages allowed. Protein identification was performed considering FDR<1% at PSM, peptide and protein level. The results from the static search were used for the statistical analysis. The protein quantification values were calculated by summing the abundance values of the peptides detected for each protein."],"omics_type":["Proteomics"],"labhead":["Fernando J. Corrales"],"instrument_platform":[""],"labhead_affiliation":["Functional Proteomics Laboratory, Centro Nacional de Biotecnologia (CSIC), Madrid 28049, Spain."],"submission_type":["COMPLETE"],"species":["Mus Musculus (mouse)"],"publication":["38488493 Blázquez-García I, Guerrero L, Cacho-Navas C, Djouder N, Millan J, Paradela A, Carmona-Rodríguez L, Corrales FJ. Molecular Insights of Cholestasis in MDR2 Knockout Murine Liver Organoids. J Proteome Res. 2024 23(4):1433-1442 10.1021/acs.jproteome.3c00900"],"submitter_mail":["ireneblazg@gmail.com"],"submitter_affiliation":["Centro Nacional de Biotecnología (CSIC)"],"submitter_country":["Spain"],"doi":["10.6019/PXD047995"],"pubmed_abstract":["MDR3 (multidrug resistance 3) deficiency in humans (MDR2 in mice) causes progressive familial intrahepatic cholestasis type 3 (PFIC3). PFIC3 is a lethal disease characterized by an early onset of intrahepatic cholestasis progressing to liver cirrhosis, a preneoplastic condition, putting individuals at risk of hepatocellular carcinoma (HCC). Hepatocyte-like organoids from MDR2-deficient mice (MDR2KO) were used in this work to study the molecular alterations caused by the deficiency of this transporter. Proteomic analysis by mass spectrometry allowed characterization of 279 proteins that were differentially expressed in MDR2KO compared with wild-type organoids. Functional enrichment analysis indicated alterations in three main cellular functions: (1) interaction with the extracellular matrix, (2) remodeling intermediary metabolism, and (3) cell proliferation and differentiation. The affected cellular processes were validated by orthogonal molecular biology techniques. Our results point to molecular mechanisms associated with PFIC3 that may drive the progression to liver cirrhosis and HCC and suggest proteins and cellular processes that could be targeted for the development of early detection strategies for these severe liver diseases."],"pubmed_title":["Molecular Insights of Cholestasis in MDR2 Knockout Murine Liver Organoids."],"pubmed_authors":["Blázquez-García Irene I, Guerrero Laura L, Cacho-Navas Cristina C, Djouder Nabil N, Millan Jaime J, Paradela Alberto A, Carmona-Rodríguez Lorena L, Corrales Fernando J FJ"],"additional_accession":[]},"is_claimable":false,"name":"Molecular insights of cholestasis in MDR2KO murine liver organoids. A model for the early detection of cirrhosis and HCC","description":"MDR3 (multidrug resistance 3) deficiency in humans (MDR2 in mice) causes progressive familial intrahepatic cholestasis type 3 (PFIC3). PFIC3 is a lethal disease characterized by an early onset of intrahepatic cholestasis, progressing to liver cirrhosis which is considered a preneoplastic condition, at risk of hepatocellular carcinoma (HCC). MDR3 is a phosphatidylcholine transporter at the canalicular membrane of hepatocytes. Hepatocyte-like organoids from MDR2-deficient mice (MDR2KO) were used in this work to study the molecular alterations caused by the deficiency of this transporter. Proteomic analysis by mass spectrometry allowed characterization of 279 proteins that were differentially expressed in MDR2KO compared to WT organoids. Functional enrichment analysis indicated alterations in three main cellular functions: (1) interaction with the extracellular matrix, (2) remodeling intermediary metabolism, (3) cell proliferation and differentiation. The affected cellular processes were validated by orthogonal molecular biology techniques such as qPCR and Western blot. Our results point to molecular mechanisms associated with PFIC3 that may drive the progression to liver cirrhosis and HCC and suggest proteins and cellular processes that could support the development of early detection strategies for these severe liver diseases.","dates":{"publication":"2026-09-17","submission":"2023-12-20"},"accession":"PXD047995","cross_references":{"TAXONOMY":["NEWT:330879","NEWT:377960","NEWT:1131","NEWT:35795","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:57577","NEWT:247480","NEWT:1211601","NEWT:876138","NEWT:44271","NEWT:193516","NEWT:2315688","NEWT:592321","NEWT:1073882","NEWT:6819","NEWT:77173","NEWT:112509","NEWT:475174","NEWT:111205","NEWT:1599","NEWT:428386","NEWT:1117","NEWT:498257","NEWT:10036","NEWT:32264","NEWT:12215","NEWT:1590","NEWT:10034","NEWT:33114","NEWT:1596","NEWT:661410","NEWT:638632","NEWT:224326","NEWT:376619","NEWT:1748418","NCBITaxon:79857","NEWT:1096976","NEWT:3765","NEWT:3767","NEWT:1589","NEWT:1907535","NEWT:135622","NEWT:4615","NEWT:67352","NEWT:672127","NEWT:35786","NEWT:1580","NEWT:399784","NEWT:96731","NEWT:656064","NEWT:1582","NEWT:3760","NEWT:1584","NEWT:2086595","NEWT:36630","NEWT:1585","NEWT:383379","NEWT:381198","NEWT:418106","NCBITaxon:12721","NEWT:10029","NEWT:913645","NEWT:2492962","NEWT:641809","NEWT:1392488","NEWT:38815","NEWT:317447","NEWT:4688","NEWT:3357","NEWT:111225","NEWT:7719","NEWT:5530","NEWT:868565","NEWT:135674","NEWT:1562698","NEWT:79329","NEWT:30069","NEWT:12637","NEWT:59729","NEWT:2164133","NEWT:295105","NEWT:29064","NEWT:108061","NEWT:60711","NEWT:224308","NEWT:273791","NEWT:3347","NEWT:49928","NEWT:3349","NEWT:160621","NEWT:2502409","NEWT:212790","NEWT:110368","NEWT:55350","NEWT:1916073","NEWT:1310161","NEWT:77133","NEWT:145481","NEWT:1310165","NEWT:29058","NCBITaxon:79824","NEWT:1912919","NEWT:410658","NEWT:5516","NEWT:44688","NEWT:44689","NEWT:370038","NEWT:498211","NEWT:345076","NEWT:347256","NEWT:5518","NEWT:398007","NEWT:1527468","NEWT:498217","NEWT:498216","NEWT:11320","NEWT:33554","NEWT:246196","NEWT:246197","NEWT:145458","NEWT:1310173","NEWT:200795","NEWT:37919","NEWT:2579791","NEWT:171719","NEWT:44685","NEWT:161934","NEWT:156587","NEWT:1148","NEWT:5508","NEWT:3329","NEWT:419481","NEWT:5507","NEWT:260499","NEWT:410661","NEWT:329854","NEWT:1140","NCBITaxon:2157","NEWT:2066563","NEWT:2649329","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:3715","NEWT:1538","NEWT:2065263","NEWT:177437","NEWT:717966","NEWT:10891","NEWT:10418","NEWT:118698","NEWT:1616117","NEWT:118696","NEWT:34865","NEWT:52283","NEWT:28174","NEWT:284812","NEWT:8175","NEWT:43330","NEWT:980415","NEWT:1603293","NEWT:44664","NEWT:43346","NEWT:3702","NEWT:66440","NEWT:1245466","NEWT:3704","NEWT:244366","NEWT:1246791","NEWT:176597","NEWT:118694","NEWT:2850","NEWT:118691","NEWT:34871","NEWT:9006","NEWT:33548","NEWT:52299","NEWT:8165","NEWT:96794","NEWT:34878","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:2714896","NEWT:1424507","NEWT:1194599","NEWT:272844","NEWT:1348799","NEWT:1608994","NEWT:33995","NEWT:55784","NEWT:1303443","NEWT:9483","NEWT:485","NEWT:56636","NEWT:2709072","NEWT:2499525","NEWT:2853422","NEWT:1122984","NEWT:487","NEWT:1538553","NEWT:1679718","NEWT:480","NEWT:67767","NEWT:45972","NEWT:46835","NEWT:109757","NEWT:582580","NEWT:1185650","NEWT:216778","NEWT:294607","NEWT:1502","NEWT:128017","NEWT:2836","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:1945864","NEWT:317047","NEWT:1509","NEWT:491","NEWT:460523","NEWT:45954","NEWT:235443","NEWT:1579","NEWT:133894","NEWT:1757312","NEWT:85694","NEWT:109786","NEWT:1922344","NEWT:108458","NEWT:5936","NEWT:85692","NEWT:321967","NEWT:320637","NEWT:1078283","NEWT:3750","NEWT:983964","NEWT:11706","NEWT:32644","NEWT:527796","NEWT:279010","NEWT:499175","NEWT:109779","NEWT:3745","NEWT:1715989","NCBITaxon:4751","NEWT:3747","NEWT:1116234","NEWT:415978","NEWT:1255228","NEWT:410289","NEWT:191701","NEWT:373153","NEWT:1640278","NEWT:9054","NEWT:472","NEWT:940825","NEWT:29459","NEWT:529507","NEWT:1071661","NEWT:28128","NEWT:470","NEWT:41122","NEWT:5911","NEWT:1328300","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:166314","NEWT:166313","NEWT:63366","NEWT:63367","NEWT:29488","NEWT:215402","NEWT:1547","NEWT:1290454","NEWT:3726","NEWT:906914","NEWT:81231006","NEWT:30455","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:299467","NEWT:9541","NEWT:8694","NEWT:33936","NEWT:8692","NEWT:542","NEWT:2903","NEWT: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