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Demeter"],"software":[""],"tissue":["Embryo","Embryonic Fibroblast"],"sample_protocol":["E1A;HrasG12V-transformed murine embryonic fibroblasts with the desired genetic modifications were plated at a density of 750,000 cells per well in a 6-well format and cultured in DMEM + 10% fetal bovine serum in an incubator held at 5% oxygen overnight. Cell lines were lysed in the plate and then protein was extracted using the iST 8x kit (Preomics P.O.00001) with the recommended protocol.   Proteins were extracted and purified form MEF cells using PreOmics iST sample preparation kit (PreOmics, P.O.00001) as instructed by the kit’s protocol. Prepared samples were dried using a SpeedVac and resuspended in Solution A (2% ACN, 0.1% FA). Sample concentrations were quantified using Quantitative Colorimetric Peptide Assay (ThermoFischer, 23275) and diluted to 100 ng/µl. 200 ng of samples were analyzed using the timsTOF Pro (Bruker Daltonics). Liquid chromatography was performed using PepSep Ten Columns (I.D. 75 µM, particle size 1.9 µm, Bruker Daltonics, 1893472) at 50ºC and peptides were separated using a 68 min gradient (solvent A: 2% ACN, 0.1% FA; solvent B: 0.1% FA in ACN) at a flow rate of 500 nl/min. A linear gradient from 3-33% B was applied for 60 min, followed by a step to 95% B for 5 min and 3 min of washing at 95% B. The timsTOF Pro was operated in PASEF mode with the following settings: Mass Range 100 to 1700m/z, 1/K0 Start 0.85 V·s/cm2, End 1.3 V·s/cm2, Ramp time 100ms, Lock Duty Cycle to 100%, Capillary Voltage 1700, Dry Gas 3 l/min, Dry Temp 200°C, PASEF settings: 4 MS/MS, charge range 0-5, active exclusion for 0.04 min, Scheduling Target intensity 20000, Intensity threshold 500, and CID collision energy 10eV."],"repository":["Pride"],"labhead":["Peter K Jackson"],"instrument_platform":[""],"labhead_affiliation":["Peter K. Jackson, Baxter Laboratory for Stem Cell Biology Department of Microbiology & Immunology Stanford University School of Medicine 269 Campus Drive Stanford, CA 94305-5175, USA"],"submission_type":["COMPLETE"],"publication":["39345444 Boutelle AM, Mabene AR, Yao D, Xu H, Wang M, Tang YJ, Lopez SS, Sinha S, Demeter J, Cheng R, Benard BA, Valente LJ, Drainas AP, Fischer M, Majeti R, Petrov DA, Jackson PK, Yang F, Winslow MM, Bassik MC, Attardi LD. Integrative multiomic approaches reveal ZMAT3 and p21 as conserved hubs in the p53 tumor suppression network. bioRxiv. 2024:2024.09.17.612743 10.1101/2024.09.17.612743"],"submitter_affiliation":["Stanford University"],"labhead_mail":["pjackson@stanford.edu"],"technology_type":["Data-dependent acquisition","Mass Spectrometry","Bottom-up proteomics"],"disease":["Disease Free"],"submitter_keywords":["Embryonic fibroblast","Mouse","P53"],"full_dataset_link":["https://www.ebi.ac.uk/pride/archive/projects/PXD047240"],"quantification_method":[""],"modification":[""],"data_protocol":["Bruker raw data files were searched against a database of NCBI mouse proteome (32,687 entries, updated 20 July 2021, only reviewed proteins) supplemented with common contaminants and reverse decoy sequences using MSFragger (version 3.5) [1] run via FragPipe (v18.0) [2]. Mass tolerances on precursors and fragments were ≤20 ppm and ≤40 ppm, respectively; protein cleavage was set to “specific” with a maximum of 2 missed cleavages per peptide. Data type was DDA, and the enzyme was trypsin, cleaving after residue K and R but not before P.  Carbamidomethylation (C) was specified as fixed while oxidation (M), acetylation (protein N-terminal) and Ser/Thr/Tyr phosphorylation were set as variable modifications. Peptide-, PTM- and ProteinProphet [3] were used for validation. Search results were then imported into Skyline (v22.1.9) [4] for ms1 quantitation using the same mass tolerances and modifications. Peak intensities were exported in MSstats format for differential analysis in R. Intensity results were summarized using MSstats [5] DataProcess function with censoredInt set to '0',  featureSubset to 'highQuality' and remove_uninformative_feature_outlier to T, with the other parameters left as default. The summarized protein level data was formatted for analysis with DEP package [6]."],"omics_type":["Proteomics"],"species":["Mus Musculus (mouse)"],"submitter_mail":["jdemeter@stanford.edu"],"submitter_country":["United States"],"doi":["10.6019/PXD047240"],"pubmed_abstract":["<i>TP53</i> , the most frequently mutated gene in human cancer, encodes a transcriptional activator that induces myriad downstream target genes. Despite the importance of p53 in tumor suppression, the specific p53 target genes important for tumor suppression remain unclear. Recent studies have identified the p53-inducible gene <i>Zmat3</i> as a critical effector of tumor suppression, but many questions remain regarding its p53-dependence, activity across contexts, and mechanism of tumor suppression alone and in cooperation with other p53-inducible genes. To address these questions, we used Tuba-seq <sup>Ultra</sup> somatic genome editing and tumor barcoding in a mouse lung adenocarcinoma model, combinatorial <i>in vivo</i> CRISPR/Cas9 screens, meta-analyses of gene expression and Cancer Dependency Map data, and integrative RNA-sequencing and shotgun proteomic analyses. We established <i>Zmat3</i> as a core component of p53-mediated tumor suppression and identified <i>Cdkn1a</i> as the most potent cooperating p53-induced gene in tumor suppression. We discovered that ZMAT3/CDKN1A serve as near-universal effectors of p53-mediated tumor suppression that regulate cell division, migration, and extracellular matrix organization. Accordingly, combined <i>Zmat3</i> - <i>Cdkn1a</i> inactivation dramatically enhanced cell proliferation and migration compared to controls, akin to <i>p53</i> inactivation. Together, our findings place <i>ZMAT3</i> and <i>CDKN1A</i> as hubs of a p53-induced gene program that opposes tumorigenesis across various cellular and genetic contexts."],"pubmed_title":["Integrative multiomic approaches reveal ZMAT3 and p21 as conserved hubs in the p53 tumor suppression network."],"pubmed_authors":["Boutelle Anthony M AM, Mabene Aicha R AR, Yao David D, Xu Haiqing H, Wang Mengxiong M, Tang Yuning J YJ, Lopez Steven S SS, Sinha Sauradeep S, Demeter Janos J, Cheng Ran R, Benard Brooks A BA, Valente Liz J LJ, Drainas Alexandros P AP, Fischer Martin M, Majeti Ravindra R, Petrov Dmitri A DA, Jackson Peter K PK, Yang Fan F, Winslow Monte M MM, Bassik Michael C MC, Attardi Laura D LD"],"additional_accession":[]},"is_claimable":false,"name":"Modeling cooperative tumor suppression in the p53 target gene network with in vivo genetic screens and multi-omics","description":"The tumor suppressor TP53 is the most frequently mutated gene in human cancer. TP53 encodes a transcriptional activator that induces numerous downstream target genes, but those genes critical for tumor suppression remain elusive. To shed light on the downstream target genes important for tumor suppression, we developed a sensitive and unbiased in vivo CRISPR/Cas9 screening platform to comprehensively map key downstream effectors of p53 in tumor suppression, an essential goal for revealing pathways that could be ultimately therapeutically targeted in p53-deficient tumors. These screens consistently revealed Zmat3 and Cdkn1a as the top-scoring tumor suppressors and demonstrated their capacity to cooperate in tumor suppression. To understand the genetic programs and cellular phenotypes regulated by Zmat3 and Cdkn1a, we used shotgun proteomics to assess differences in protein expression between wild-type, Zmat3 KO, Zmat3 + Cdkn1a KO, and p53 KO E1A;HrasG12V murine embryonic fibroblasts. Functional annotation revealed that various cellular programs including cell division, extracellular signaling and signal transduction, viral response, and ribosome biogenesis become dysregulated in cell lines with alterations to the p53 pathway compared to the wild-type cell lines. These data suggest cellular processes and signaling pathways that provide clues to understanding p53-mediated tumor suppression and for the development of treatments for p53 deficient tumors.","dates":{"publication":"2026-07-17","submission":"2023-11-25"},"accession":"PXD047240","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:247480","NEWT:1211601","NEWT:876138","NEWT:44271","NEWT:193516","NEWT:2315688","NEWT:1073882","NEWT:6819","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","NCBITaxon:79857","NEWT:1096976","NEWT:3765","NEWT:3767","NEWT:1589","NEWT:135622","NEWT:4615","NEWT:672127","NEWT:35786","NEWT:1580","NEWT:399784","NEWT:96731","NEWT:656064","NEWT:1582","NEWT:3760","NEWT:1584","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: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:3349","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:345076","NEWT:347256","NEWT:5518","NEWT:398007","NEWT:1527468","NEWT:498217","NEWT:498216","NEWT:11320","NEWT:246196","NEWT:246197","NEWT:145458","NEWT:1310173","NEWT:200795","NEWT:2579791","NEWT:44685","NEWT:161934","NEWT:156587","NEWT:1148","NEWT:5508","NEWT:3329","NEWT:419481","NEWT:5507","NEWT:260499","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:3715","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: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:9006","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: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:317047","NEWT:1509","NEWT:491","NEWT:460523","NEWT:45954","NEWT:235443","NEWT:1579","NEWT:1757312","NEWT:85694","NEWT:109786","NEWT:108458","NEWT:5936","NEWT:321967","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:415978","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:41122","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:29488","NEWT:215402","NEWT:1547","NEWT:1290454","NEWT:3726","NEWT:906914","NEWT:81231006","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:542","NEWT:2903","NEWT:185579","NEWT:872227","NEWT:13076","NEWT:2587410","NEWT:207340","NEWT:1006581","NEWT:8673","NEWT:33940","NEWT:2126337","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:64657","NEWT:186441","NEWT:643680","NEWT:214092","NCBITaxon:6157","NEWT:13095","NEWT:162425","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