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To induce ER stress, cells were treated with 5 µg/mL tunicamycin (TM) for 4 hours, and DMSO at 1:1000 dilution was used as a control. Cells were washed with ice-cold PBS and collected by scraping and pelleting at 500 g at 4°C for 5 min. Cells were lysed in 250 µL of lysis buffer (25 mM HEPES pH 7.3, 150 mM NaCl, 0.5% NP-40, 0.5 mM EDTA, 10% Glycerol, 1x EDTA-free protease inhibitor cocktail, 1x PhosSTOP) per condition by incubation on ice 15 min with intermittent vortexing. The lysate was clarified by centrifugation at 20,000 g for 20 min at 4°C. 10 μg of Proteintech anti-IGF2BP3 antibody (14642-1-AP, lot 00088732) or Proteintech IgG control (30000-0-AP) was coupled to the 40 μL of protein G Dynabeads (Invitrogen) in 1 mL of lysis buffer for 20 min, rotating at room temperature, washed three times with 1 mL of the lysis buffer, resuspended in the original bead volume (40 μL) and added to 200 µL of the clarified lysate. The IP was incubated on a rotator at +4°C for 3 h. Beads were washed five times with 1 mL of wash buffer (25 mM HEPES pH 7.3, 150 mM NaCl, 0.5 mM EDTA, 10% Glycerol). Each wash was incubated on ice for 3 min. During the second wash “RNase” samples were incubated in 250ul ul of wash buffer + 1:1000 RNase I Ambion + 1:1000 RNase T1 (Thermo, EN0541) for 10 min rotating at room temperature. For the second wash “noRNase” samples were kept on ice in in 250ul ul of wash buffer. The rest of the washes were performed as described above. Samples were submitted for on-bead tryptic digestion and mass spectrometry. The proteins were eluted from the beads with 20µL 100mM Glycine pH2 3 times and the pooled supernatant was adjusted to alkaline using about 20µL 1M Tris pH 8. Disulfide bonds were reduced with 3.2 µL of 250 mM dithiothreitol (DTT) for 30 min at room temperature before adding 3.2 µL of 500 mM iodoacetamide and incubating for 30 min at room temperature in the dark. Remaining iodoacetamide was quenched with 1.6 µL of 250 mM DTT for 10 min. Proteins were digested with 300 ng trypsin (Trypsin Gold, Promega) in 3 µL 50 mM ammonium bicarbonate at 37°C over night. The digest was stopped by the addition of 10% trifluoroacetic acid (TFA) to a final concentration of 0.5%. The peptides were desalted using C18 Stagetips. Peptides were separated on a Vanquish Neo nano-flow chromatography system (Thermo-Fisher), using a trap-elute method for sample loading (Acclaim PepMap C18, 2 cm × 0.1 mm, 5 μm, Thermo-Fisher), and a C18 analytical column (Acclaim PepMap C18, 50 cm × 0.075 mm, 2 μm, Thermo-Fisher), applying a segmented linear gradient from 2% to 35% and finally 80% solvent B (80 % acetonitrile, 0.1 % formic acid; solvent A 0.1 % formic acid) at a flow rate of 230 nL/min over 120min. Eluting peptides were analyzed on an Exploris 480 Orbitrap mass spectrometer (Thermo-Fisher Scientific) coupled to the column with a FAIMS pro ion-source (Thermo-Fisher Scientific) using coated emitter tips (PepSep, MSWil) with the following settings: The mass spectrometer was operated in DDA mode with two FAIMS compensation voltages (CV) set to -45 or -60 and 1.5 s cycle time per CV. The survey scans were obtained in a mass range of 350-1500 m/z, at a resolution of 60k at 200 m/z, and a normalized AGC target at 100%. The most intense ions were selected with an isolation width of 1.4 m/z, fragmented in the HCD cell at 30% collision energy, and the spectra recorded for max. 50 ms at a normalized AGC target of 100% and a resolution of 15k. Peptides with a charge of +2 to +6 were included for fragmentation, the peptide match feature was set to preferred, the exclude isotope feature was enabled, and selected precursors were dynamically excluded from repeated sampling for 45 seconds."],"repository":["Pride"],"quantification_method":["Not available"],"modification":[""],"data_protocol":["The RAW MS data were analyzed with FragPipe (22.0), using MSFragger (4.1), IonQuant (1.10.27) and Philosopher (5.0.0). The default FragPipe workflow for label free quantification (LFQ-MBR) was used, except “Normalize intensity across runs” was turned off. Cleavage specificity was set to Trypsin/P, with two missed cleavages allowed. The protein FDR was set to 1%. A mass of 57.02146 (carbamidomethyl) was used as fixed cysteine modification; methionine oxidation and protein N-terminal acetylation were specified as variable modifications. MS2 spectra were searched against the human 1 protein per gene reference proteome from Uniprot (Proteome ID: UP000005640, release 2024_01), concatenated with a database of 382 common laboratory contaminants (release 2023.03, https://github.com/maxperutzlabs-ms/perutz-ms-contaminants). Computational analysis was performed using Python and the in-house developed Python library MsReport (version 0.0.24,). LFQ protein intensities reported by FragPipe were log2-transformed and normalized across samples using the ModeNormalizer from MsReport. The missing normalized LFQ intensity values were imputed by drawing random values from a normal distribution after filtering out contaminants, proteins with less than 2 peptides and less than 2 quantified values in at least one group. Differences between groups were statistically evaluated using the LIMMA 3.52.1 at 5% FDR (Benjamini-Hochberg). The in-house Python library XlsxReport (0.1.0) was used to create a formatted Excel file summarizing the results of protein quantification."],"omics_type":["Proteomics"],"labhead":["Dr Gülsün Elif Karagöz"],"instrument_platform":[""],"labhead_affiliation":["Max Perutz Labs, Vienna BioCenter Campus (VBC), 1030 Vienna, Austria; Austria Medical University of Vienna, Center for Medical Biochemistry, 1030 Vienna, Austria"],"submission_type":["PARTIAL"],"species":["Homo Sapiens (human)"],"submitter_mail":["proteomics@maxperutzlabs.ac.at"],"publication":["10.1101/GAD.353291.125"],"submitter_affiliation":["Max Perutz Labs, Mass Spectrometry Facility, Vienna Biocenter Campus (VBC), Dr. Bohr-Gasse 9, Vienna, Austria"],"submitter_country":["Austria"],"additional_accession":[]},"is_claimable":false,"name":"Identification of RNA-mediated interaction partners of endogenous IGF2BP3 in HCT116 cells in control and ER stress conditions","description":"IGF2BP3 is an RNA-binding protein that regulates mRNA stability, translation, and localization. IGF2BP3 binds to mRNAs encoding endoplasmic reticulum (ER) stress response proteins. To understand molecular mechanisms of its function we performed co-immunoprecipitation of the endogenous IGF2BP3 from colorectal carcinoma cell line HCT116 in control and ER stress conditions. RNase treatment was performed during washes to reveal RNA-bridged interactors.","dates":{"publication":"2026-07-24","submission":"2025-02-06"},"accession":"PXD060548","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:1211601","NEWT:876138","NEWT:44271","NEWT:193516","NEWT:2315688","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:4615","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",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