{"database":"Pride","file_versions":[{"headers":{"Content-Type":["application/json"]},"body":{"files":{"Txt":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/checksum.txt","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/metadata_Zip1.txt","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/evidence.txt"],"Xml":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/mqpar.xml"],"Raw":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010368.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010366.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010364.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010359.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010360.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010361.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010363.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010357.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010367.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010358.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010365.raw","ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/EX010362.raw"],"Fasta":["ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/08/PXD075416/UP000000625_EColi_Zip1_correncted_w_cRAP_contaminants.fasta"]},"type":"primary"},"statusCode":"OK","statusCodeValue":200}],"scores":null,"additional":{"labhead_mail":["pitter.huesgen@biologie.uni-freiburg.de"],"submitter":["Pitter Huesgen"],"technology_type":["Data-dependent acquisition","Mass Spectrometry","Bottom-up proteomics"],"software":[""],"submitter_keywords":["Apoplastic immunity","Phytocytokine","Plcps","Zip1","Metacaspase"],"full_dataset_link":["https://www.ebi.ac.uk/pride/archive/projects/PXD075416"],"sample_protocol":["Recombinant PROZIP1, PROZIP1CS and PROZIP1noR and were incubated with either recombinant active or inactive ZmMC9 or ZmMC9C141A for 10 min and 20 min- Proteolysis was stopped by the addition of 10 mM EDTA and 500 µM leupeptin before dimethyl labeling with 40 mM formaldehyde (13CH₂O) and 40 mM sodium cyanoborohydrate, precipitated Sera-Mag SpeedBead magnetic carboxylate-modified particles (Cytiva) before on-bead digestion with trypsin or legumain. Peptides were eluted from the beads, dried using a SpeedVac concentrator and reconstituted in 0.1% formic acid prior to loading onto Evotips (Evotip Pure EV2013, Evosep).  Stage-tip purified peptides were separated by nanoflow HPLC (Thermo Ultimate RSLCnano equipped with Thermo Acclaim or PharmaFluidics PAC trap and analytical columns), using a 2–80% ACN gradient, and analyzed with a Bruker QqTOF (Impact II) using a CaptiveSpray ion source. MS1 scans (m/z 150–3000, 2 Hz) were followed by CID of the Top14 precursors with 45 s dynamic exclusion. Peptides loaded on Evotips in the MC9 experiment were analysed with an Evosep One system (Evosep) coupled to an Exploris 480 Orbitrap mass spectrometer (Thermo) with a Nanospray Flex ion source equipped with a stainless-steel emitter with integrated liquid junction (EV1072) mounted in an EasySpray adapter. Peptides were separated using a 21-min gradient (60 samples per day workflow) on an 8 cm × 150 µm Evosep performance column (EV1137). MS1 survey scan (m/z 350–1400) acquired at 120,000 resolution in profile mode, followed by HCD fragmentation of the 12 most intense precursors with 30s dynamic exclusion."],"repository":["Pride"],"quantification_method":["Not available"],"modification":[""],"data_protocol":["Acquired mass spectrometry data were analyzed with MaxQuant (v2.4.20) with standard instrument settings using the E. coli protein databases with appended recombinant PROZIP1 sequences and ZmMC9 sequences in the appropriate combination for each experiment. Digestion mode was set to semi-specific (in vivo, MC9), with dimethylation (N-term, Lys), methionine oxidation, and N-terminal acetylation as variable modifications."],"omics_type":["Proteomics"],"labhead":["Pitter F Huesgen"],"instrument_platform":[""],"labhead_affiliation":["Institute of Biology II, University of Freiburg, Freiburg, Germany Central Institute for Engineering, Electronics and Analytics, ZEA-3, Forschungzentrum Jülich, Jülich, Germany CIBSS-Centre for Integrative Biological Signaling Studies, University of Freiburg, Freiburg, Germany"],"submission_type":["PARTIAL"],"species":["Escherichia Coli"],"submitter_mail":["pitter.huesgen@biologie.uni-freiburg.de"],"publication":["42483934 Koenig M, Sorger Z, Kakanj P, Dewes P, Mantz M, Perrar A, Sivaramakrishnan M, Stael S, Chandrasekar B, Huesgen PF, Villamil JM, Doehlemann G. Processing and release of the maize phytocytokine Zip1. Plant Physiol. 2026:kiag533 10.1093/plphys/kiag533"],"submitter_affiliation":["University of Freiburg\nFaculty of Biology\nBiochemistry and Functional Proteomics"],"submitter_country":["Germany"],"pubmed_abstract":["Phytocytokines are endogenous peptides that modulate plant immunity outcomes, yet how their maturation and spatial deployment are controlled remains unclear. Here we show that the maize phytocytokine precursor PROZIP1 is controlled by a spatially separated, two-stage proteolytic pathway that mechanistically uncouples signal activation from extracellular attenuation. PROZIP1 associates with the endoplasmic reticulum and undergoes intracellular, arginine-dependent processing by type II metacaspases, generating a C-terminal PROZIP1 fragment (Ct-PROZIP1). This processing licenses PROZIP1 for export to the apoplast via an ER-Golgi-independent route. Proteomic mapping and mutational analyses identify arginine residues flanking the Zip1 peptide as critical for efficient processing and secretion. The calcium-dependent metacaspase ZmMC9 specifically processes PROZIP1, thereby efficiently generating the bioactive Ct-PROZIP1 fragment. In the apoplast, Ct-PROZIP1 is further processed by papain-like cysteine proteases and additional extracellular proteases, contributing to Zip1 turnover and signal clearance. While the free Zip1 peptide is detected at later stages, Ct-PROZIP appears to be the primary signaling entity in modulating pathogen-induced immune responses. Together, these findings demonstrate a previously unknown complexity in peptide signaling, suggesting a multilayered control of phytocytokine activity that provides spatial and temporal precision to disease modulation in maize."],"pubmed_title":["Processing and release of the maize phytocytokine Zip1."],"pubmed_authors":["Koenig Maurice M, Sorger Zarah Z, Kakanj Parisa P, Dewes Paula P, Mantz Melissa M, Perrar Andreas A, Sivaramakrishnan Muthusaravanan M, Stael Simon S, Chandrasekar Balakumaran B, Huesgen Pitter F PF, Villamil Johana Misas JM, Doehlemann Gunther G"],"additional_accession":[]},"is_claimable":false,"name":"Processing and release of the maize phytocytokine Zip1","description":"Phytocytokines are endogenous peptides that modulate plant immunity outcomes, yet how their maturation and spatial deployment are controlled remains unclear. Here we show that the maize phytocytokine precursor PROZIP1 is controlled by a spatially separated, two-stage proteolytic pathway that mechanistically uncouples signal activation from extracellular attenuation. PROZIP1 associates with the endoplasmic reticulum and undergoes intracellular, arginine-dependent processing by type II metacaspases, generating a C-terminal PROZIP1 fragment (Ct-PROZIP1). This processing licenses PROZIP1 for export to the apoplast via an ER-Golgi-independent route. Proteomic mapping and mutational analyses identify arginine residues flanking the Zip1 peptide as critical for efficient processing and secretion. The calcium-dependent metacaspase ZmMC9 specifically processes PROZIP1, thereby efficiently generating the bioactive Ct PROZIP1 fragment. In the apoplast, Ct-PROZIP1 is further processed by papain-like cysteine proteases and additional extracellular proteases, contributing to Zip1 turnover and signal clearance. While the free Zip1 peptide is detected at later stages, Ct-PROZIP appears to be the primary signaling entity in modulating pathogen-induced immune responses. Together, these findings demonstrate a previously unknown complexity in peptide signaling, suggesting a multilayered control of phytocytokine activity that provides spatial and temporal precision to disease modulation in maize.","dates":{"publication":"2026-08-04","submission":"2026-03-09"},"accession":"PXD075416","cross_references":{"TAXONOMY":["NEWT:6945","NEWT:3555","NEWT:241368","NEWT:2","NEWT:157546","NEWT:190802","NEWT:35554","NEWT:9778","NEWT:150475","NEWT:9417","NEWT:347515","NEWT:1216979","NEWT:307972","NEWT:32046","NEWT:544496","NEWT:5180","NEWT:256737","NEWT:2042546","NEWT:115104","NEWT:1081927","NEWT:67825","NEWT:43179","NEWT:13076","NEWT:1249668","NEWT:376741","NEWT:317","NEWT:55153","NEWT:1736309","NEWT:7227","NEWT:7469","NEWT:885318","NEWT:415540","NEWT:876138","NEWT:4081","NEWT:554","NEWT:98334","NEWT:426428","NEWT:237561","NEWT:6928","NEWT:10036","NEWT:7574","NEWT:1351","NEWT:7215","NEWT:29204","NEWT:272563","NEWT:507601","NCBITaxon:79857","NCBITaxon:6157","NEWT:95648","NEWT:3885","NEWT:746360","NEWT:6239","NEWT:1589","NEWT:470150","NEWT:135622","NEWT:216257","NEWT:6915","NEWT:9986","NEWT:101510","NEWT:4054","NEWT:3880","NEWT:3641","NEWT:8782","NEWT:1263854","NEWT:1000589","NEWT:1902","NEWT:85962","NEWT:160488","NEWT:28104","NEWT:317447","NEWT:7955","NCBITaxon:2","NEWT:985076","NEWT:7959","NEWT:2261","NEWT:4565","NEWT:1264690","NEWT:6192","NEWT:28532","NCBITaxon:38727","NEWT:34305","NEWT:59729","NCBITaxon:183674","NEWT:224308","NEWT:626528","NEWT:139927","NEWT:4558","NEWT:209285","NEWT:216595","NEWT:243230","NEWT:8355","NEWT:1283","NEWT:931281","NEWT:1000561","NEWT:7029","NEWT:1283300","NEWT:6183","NEWT:334747","NEWT:61235","NCBITaxon:79824","NEWT:4787","NCBITaxon:4563","NEWT:5755","NEWT:3218","NEWT:5759","NEWT:1736231","NEWT:436486","NEWT:6287","NEWT:2242","NEWT:300641","NEWT:4784","NEWT:727","NEWT:9796","NEWT:725","NEWT:360106","NEWT:260707","NEWT:287","NEWT:10117","NEWT:10239","NCBITaxon:6191","NEWT:10116","NEWT:1280","NEWT:1836","NEWT:1735272","NEWT:83334","NEWT:185431","NEWT:83332","NEWT:29760","NEWT:260704","NEWT:703612","NEWT:260705","NEWT:80863","NEWT:44685","NEWT:2697049","NEWT:1148","NEWT:11676","NEWT:55571","NEWT:100226","NCBITaxon:6073","NEWT:4530","NEWT:4896","NEWT:6279","NEWT:1123869","NEWT:7370","NEWT:75058","NEWT:83906","NEWT:607699","NEWT:6282","NEWT:208964","NEWT:1134506","NEWT:575584","NEWT:296543","NEWT:1773","NEWT:38783","NEWT:8727","NEWT:1895","NEWT:1182590","NEWT:8726","NEWT:6669","NEWT:10090","NEWT:935293","NEWT:64152","NEWT:749200","NEWT:4120","NEWT:51515","NEWT:5693","NEWT:8724","NEWT:51511","NEWT:92867","NEWT:8723","NEWT:990346","NEWT:5334","NEWT:145953","NEWT:257309","NEWT:100816","NEWT:230741","NEWT:284812","NCBITaxon:10359","NCBITaxon:1313","NEWT:43330","NEWT:242619","NEWT:44544","NEWT:632957","NEWT:373995","NEWT:5689","NEWT:645463","NEWT:544404","NEWT:3702","NEWT:129249","NEWT:9925","NEWT:8839","NEWT:4232","NEWT:990119","NEWT:2758385","NEWT:4113","NEWT:837","NEWT:11298","NEWT:171101","NEWT:421932","NEWT:196627","NEWT:408172","NEWT:5691","NEWT:408170","NEWT:493760","NEWT:260710","NEWT:627025","NEWT:400772","NEWT:1097677","NEWT:3708","NEWT:128161","NEWT:106592","NEWT:1117957","NEWT:9913","NEWT:1432138","NEWT:10312","NEWT:1424507","NEWT:4100","NEWT:1076","NEWT:6763","NEWT:803","NEWT:8030","NEWT:29722","NEWT:380394","NEWT:1692259","NEWT:1639","NEWT:188229","NEWT:3818","NEWT:480","NEWT:4909","NEWT:180066","NEWT:67767","NEWT:46835","NEWT:135588","NEWT:1843183","NEWT:95486","NEWT:58002","NEWT:9103","NEWT:4577","NEWT:5664","NEWT:2157","NEWT:146479","NEWT:10306","NEWT:1911079","NEWT:8022","NEWT:145943","NCBITaxon:4932","NEWT:595536","NEWT:3635","NEWT:5811","NEWT:235443","NEWT:1480154","NEWT:1274414","NEWT:59202","NEWT:9975","NEWT:3197","NEWT:9615","NEWT:10299","NEWT:860688","NEWT:884019","NEWT:169963","NEWT:36329","NEWT:1147787","NEWT:72407","NEWT:9606","NEWT:367830","NEWT:157295","NEWT:178616","NEWT:410289","NEWT:373153","NEWT:915099","NEWT:74940","NEWT:1450511","NEWT:360094","NEWT:470","NEWT:84023","NEWT:9838","NCBITaxon:9615","NEWT:58334","NEWT:1193501","NEWT:3055","NEWT:6326","NEWT:6689","NEWT:2762","NEWT:5476","NEWT:1174673","NEWT:562","NEWT:33952","NEWT:1274432","NEWT:1274426","NEWT:1423","NEWT:4932","NEWT:70448","NEWT:9825","NEWT:1274423","NEWT:3603","NEWT:698936","NEWT:2759","NEWT:3847","NEWT:39946","NEWT:9823","NEWT:9940","NEWT:327160","NEWT:573","NEWT:9031","NEWT:1274420","NEWT:7091"],"pubmed":["42483934"],"ORCID":["0000-0002-0335-2242"]}}