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TholenMass SpectrometryShotgun proteomicsNot availableDegradomicsProteomicsSkinhttp://www.ebi.ac.uk/pride/archive/projects/PXD002597SkinlProtein lysates the epidermis was detached from the dermis by heat separation. Epidermis was lysed in RIPA buffer. Proteins were precipitated using acetone and digested using trypsin. Formaldehyde labeling was performed as a label-switch experiment, meaning the wild type sample was labeled light in the first biological replicate and heavy in the second biological replicate. Labeling was performed after tryptic digestion. Data were converted to mzXML format using Proteowizard with centroiding of MS1 and MS2 data. Peptide sequences were identified by X! Tandem (version 2013.09.01.1), including cyclic permutation, in conjunction with PeptideProphet (part of version 4.7 of the Trans Proteomic Pipeline (TPP)) and a decoy search strategy: the complete mouse proteome file was downloaded from UniProt on Nov 26th 2013. It was appended with an equal number of randomized sequences, derived from the original mouse proteome entries. The decoy database was generated with DBToolkit. Tryptic cleavage specificity with no missed cleavage sites was applied. Mass tolerance was 10 ppm for parent ions and 0.3 Da for fragment ions. Static modifications are cysteine carboxyamidomethylation (+57.02 Da), lysine and N-terminal dimethylation (light formaldehyde 28.03 Da; heavy formaldehyde 34.06 Da). X!Tandem results were further validated by PeptideProphet at a confidence level of > 95 %. Corresponding protein identifications are based on the ProteinProphet algorithm with a protein false discovery rate of < 1 %. The relative quantitation for each protein was calculated from the relative areas of the extracted ion chromatograms of the precursor ions and their isotopically distinct equivalents using the XPRESS algorithm. Reported Fc values are based on normalized XPRESS ratios. Proteins were considered to be altered in their abundance if they were (A) identified in both biological replicates of each experimental setup, (B) showed an alteration in abundance of more than 50 % (Fc < -0.58; Fc > 0.58) and (C) if their H/L ratio calculated by XPRESS was confirmed by manual inspection of the extracted ions chromatograms. TAILS was performed using formaldehyde labeling according to the original publication [22]. 2 mg of protein were used per condition. Formaldehyde labeling was performed as a label-switch experiment, meaning the wild type sample was labeled light in the first biological replicate and heavy in the second biological replicate. After tryptic digest samples were desalted using a reversed phase C18 column, prefractionated by SCX as described, and desalted using self-packed C18 STAGE tips (Empore, USA) [33]. LC-MS/MS analysis is described in the corresponding section. As for the quantitative proteome comparison, data were converted to mzXML format [26] using Proteowizard [27] with centroiding of MS1 and MS2 data. Peptide sequences were identified by X! Tandem (version 2013.09.01.1) [28], including cyclic permutation, in conjunction with PeptideProphet (part of version 4.7 of the Trans Proteomic Pipeline) [29] and a decoy search strategy: the complete mouse proteome file was downloaded from UniProt [30] on Nov 26th 2013. Semi Arg-C specificity with up to two missed cleavage sites was applied. Static modifications are (+ 57.02 Da), lysine and N-terminal dimethylation (light formaldehyde + 28.03 Da; heavy formaldehyde + 34.06 Da). For acetylated N-termini (+ 42.01 Da), modifications are cysteine carboxyamidomethylation, N-terminal acetylation, and lysine dimethylation. Mass tolerance was 10 ppm for parent ions and 0.02 Da for fragment ions. X!Tandem results were further validated by PeptideProphet at a confidence level of > 95 %. The relative quantification for each peptide was calculated using the XPRESS [32] algorithm as described for “Quantitative Proteome Comparison”. Two biological replicates comparing wild type and ADAM17ΔKC epidermal lysates of littermates at P3 with independent sample preparation (not the same samples as for quantitative proteome comparison) and mass spectrometric measurement were analyzed. N-termini were considered to be altered in their abundance if they were (A) identified in both biological replicates, (B) showed an alteration in abundance of more than 50 % (Fc < -0.58; Fc > 0.58) and (C) if their H/L ratio calculated by XPRESS was confirmed by manual inspection of the extracted ions chromatograms (XIC).PrideNot availableNo PTMs are included in the datasetLC-MS/MS Analysis For nanoflow-LC-MS/MS, MS samples were analyzed on an Orbitrap XL (Thermo Scientific GmbH, Bremen, Germany) mass spectrometer. The instrument was coupled to an Ultimate3000 micro pump (Thermo Scientific) with a flow rate or 300 nl / min. 0.5 % acetic acid and 0.5 % acetic acid in 80 % acetonitrile (water and acetonitrile were at least HPLC gradient grade quality) with a gradient of increasing organic proportion were used for peptide separation. Column-tips with 75 µm inner diameter and a length of 11 cm were self-packed [34] with Reprosil-Pur 120 ODS-3 (Dr. Maisch, Ammerbuch, Germany). The mass spectrometer was operated in the data dependent mode and switched automatically between MS and MS/MS.ProteomicsOliver SchillingLTQ OrbitrapInstitute of Molecular Medicine and Cell Research University of Freiburg Stefan Meier Strasse 17 79104 Freiburg, GermanyPARTIALMus Musculus (mouse)stefan.tholen@mol-med.uni-freiburg.de27089454 Tholen S, Wolf C, Mayer B, Knopf JD, Löffek S, Qian Y, Kizhakkedathu JN, Biniossek ML, Franzke CW, Schilling O. The Skin Barrier Defects Caused by Keratinocyte-Specific Deletion of ADAM17 or EGFR are Based on Highly Similar Proteome and Degradome Alterations. J Proteome Res. 2016 Apr 11BiologicalBiomedicalInstitute for Molecular MedicineKeratinocyte-specific deletion of ADAM17 in mice impairs terminal differentiation of keratinocytes leading to severe epidermal barrier defects. Mice deficient for ADAM17 in keratinocytes phenocopy mice with a keratinocyte-specific deletion of epidermal growth factor receptor (EGFR), which highlights the role of ADAM17 as a "ligand sheddase" of EGFR ligands. In this study, we aim for the first proteomic/degradomic approach to characterize the disruption of the ADAM17-EGFR signaling axis and its consequences for epidermal barrier formation. Proteomic profiling of the epidermal proteome of mice deficient for either ADAM17 or EGFR in keratinocytes at postnatal days 3 and 10 revealed highly similar protein alterations for ADAM17 and EGFR deficiency. These include massive proteome alterations of structural and regulatory components important for barrier formation such as transglutaminases, involucrin, filaggrin, and filaggrin-2. Cleavage site analysis using terminal amine isotopic labeling of substrates revealed increased proteolytic processing of S100 fused-type proteins including filaggrin-2. Alterations in proteolytic processing are supported by altered abundance of numerous proteases upon keratinocyte-specific Adam17 or Egfr deletion, among them kallikreins, cathepsins, and their inhibitors. This study highlights the essential role of proteolytic processing for maintenance of a functional epidermal barrier. Furthermore, it suggests that most defects in formation of the postnatal epidermal barrier upon keratinocyte-specific ADAM17 deletion are mediated via EGFR.Skin Barrier Defects Caused by Keratinocyte-Specific Deletion of ADAM17 or EGFR Are Based on Highly Similar Proteome and Degradome Alterations.Tholen Stefan S, Wolf Cristina C, Mayer Bettina B, Knopf Julia D JD, Löffek Stefanie S, Qian Yawen Y, Kizhakkedathu Jayachandran N JN, Biniossek Martin L ML, Franzke Claus-Werner CW, Schilling Oliver O482DER/faint little ball, HSN1E, second cervical vertebra, determination, Laboratory, Mus domesticus, Glutaminyl-Peptide Gamma-Glutamyltransferases, Elp-B1, der, DER1., Elp-1, cleavage, A4, Errb1, CASP-14, protein, House Mouse, 10-dimethyl-3-(1-methylethyl)-11-oxabicyclo(8.1.0)undec-6-en-2-yl ester, binding site, PIG61, axis vertebra, protein polypeptide chains, Roles, kinky, epidermal growth factor-activated receptor activity, AI552599, Concepts, 3, 4, d-egf-r, protein aggregate, multicellular organismal biosynthetic process, AI266795, increased, single-organism biosynthetic process, activation inducer molecule, anabolism, Kininogenase, C2, Swiss Mice, proteins, NISBD, flb, Protein Glutamine gamma Glutamyltransferases, vertebra 2, signaling process, Role Concepts, AI316800, keratinized cell of epidermis, Protein-Glutamine, NISBD1, 9030024J15Rik, D-EGFR, fused to, Kallikrein Light Chain, beta Kallikrein, NISBD2, single organism signaling, DER1, Ligand, Elp, Wa5, l(2)09261, DEgfr, mouse, beta-Kallikrein, C2 vertebra, S100, S100-B, CG10079, torpedo/egfr, Role Concept, Kallikrein B', wa2, Callicrein, Mini-ICE, Glutaminyl Peptide Gamma Glutamyltransferases, Kallidinogenase, Role, region, Kallikrein A, egfr, adam18, Mus musculus, early T-cell activation antigen p60, ADCADN, HD-33, El, positional polypeptide feature, Caspase-14 subunit p10, 5-trimethoxy-, mice, EGFR, DER/EGFR, EGfr, EgfR, FU, dEgfr, Swiss Mouse, DER flb, Caspase-14 subunit p19, gamma-Glutamyltransferases, Torpedo/Egfr, axis [C II], MICE, 4-(acetyloxy)-6, merged with, domesticus, Degfr, DMDA, Kalliginogenase, DEGFR, l(2)57DEFa, l(2)05351, Protein-Glutamine gamma-Glutamyltransferases, Fu, beta-Kallikrein B, dEGFR, Mouse, DmHD-33, fused, CD69, Proteomes, accessory, Benzoic acid, FUSED, 3.4.22.-, receptor tyrosine-protein kinase erbB-1, knobbly, malpighian cell, csvp, biological signaling, D-Egf, proto-oncogene c-ErbB-1, Transglutaminase, Filaggrin, region or site annotation, Protein-glutamine:amine gamma-glutamyltransferase, alpha-Kallikrein, C-erb, mor1, Fused, EA1, Gene, mini-ICE, stalk, Gamma-Glutamyltransferases, biosynthesis, mENA, MLR-3, EK2-6, protein-containing complex, terminal differentiation, supernumerary, TYPE, DAGA4, Light Chain, AIM, polypeptide chain, House, c-erbB, EGFr, Egfr, S100-alpha, Gene Products, ADAM18, Mus musculus domesticus, EGF receptor activity, torpedo/Egfr, TOP, CSVP, Egf-r, MAM, early activation antigen CD69, Mice, SCG3, study, EGF-R, positional, formation, Swiss, Dilminal, ligand, axis (CII), top, Torpedo/DER, labeling, Maintenances, culm, top/flb, synthesis, S100beta, alpha Kallikrein, leukocyte surface antigen Leu-23, Enzyme, Stratum Corneum Basic Protein Precursor, Kallikrein, l(2)57EFa, NEF, C-type lectin domain family 2 member C, site, Kinin Forming Enzyme, l(2)57Ea, mKIAA1278, Kb, cd156b, Ki, Elp-B1RB1, Filaggrin Protein, Profilaggrin, Tace, TGF-alpha receptor activity, axis, protein complex, ERBB, dEGFR1, Proteins, DNMT, Stratum Corneum Basic Protein, total expressed protein, MCMT, B930045J24, LGMD2C, Concept, BL-AC/P26, Axin, cervical axis, TACE, epidermal growth factor receptor activity, native protein, DmelCG10079, natural protein, Mus, Protein, chemical analysis, DER/top, tace, CXXC9, Padutin, DMDA1, top/DER, S100a, INSDC_feature:misc_binding, increased number, cervical vertebra 2, Cathepsin, Amine, beta Kallikrein B, House Mice, ERBB1, wa-2, Keratinocyte, CD156b, signalling, Laboratory Mice, Protein Gene Products, present in greater numbers in organism, Gene Proteins, Egf, EFG-R, signalling process, binding_or_interaction_site, GP32/28, DEL, Errp, S100A, Erbb, joined with, Glutaminyl-Peptide, SCARMD2, Der, DER, transforming growth factor-alpha receptor activity, assay, 1700112N14Rik, CD156B, Laboratory Mouse, coalesced, DER/torpedo, CLEC2C, HER1, Kinin-Forming, Kinin-Forming Enzymeliquid chromatography tandem mass spectroscopy, LC-MS2, DYRK1, AcOH, E-260, MeCN, NCMe, 1-(14)C-labeled, determination, 3H-labeled, Vinegar, instrument configuration, LC-MS/MS, Dmel_CG7826, CH3-COOH, Essigsaeure, acetonitrile, Thermo Scientific, Peptide, CG7826, LC-MS-MS, polypeptide, peptide, Polypeptides, Methanecarboxylic acid, LC/MS/MS, Glacial, E260, HOAc, ClvPrd, peptido, Dm1, Glacial Acetic Acid, CH3-C#N, LC-MSMS, chemical analysis, CG7835, CG42273, Min, Dyrk1, ACETONITRILE, Dmel_CG7835, CH3CO2H, Acetic acid, DmelCG42273, cyanomethane, Hydrogen Oxide, Mnb, MNB, AU020952, proportion, LCMSMS, Ethylic acid, peptides, instrument, E 260, CC1, proportionality, Acetic Acid, min, mAPC, rate, acide acetique, ethanenitrile, AW124434, Acetic Acid Glacial, HPLC, ME-IV, AI047805, 10^[-6], column, MeCOOH, ACETIC ACID, liquid chromatography-tandem mass spectroscopy, data., MeCO2H, quotient, liquid chromatography tandem mass spectrometry, ethoic acid, INS No. 260, Peptid, peptidos, assay, Polypeptide, methyl cyanide, acetic acid, Ethanoic acid, ratioDER/faint little ball, dermis plus epidermis plus hypodermis, receptor tyrosine-protein kinase erbB-1, the integument, malpighian cell, csvp, region of skin, D-Egf, pelt, proto-oncogene c-ErbB-1, Otf-11, Skn-li, C-erb, Elp-B1, der, Elp-1, mor1, Errb1, mENA, EK2-6, epidermis, PIG61, epidermal growth factor-activated receptor activity, AI552599, c-erbB, EGFr, Egfr, ADAM18, EGF receptor activity, torpedo/Egfr, TOP, CSVP, Egf-r, d-egf-r, integumentum commune, Skn-1a, EGF-R, skin, dermoid system, entire skin, top, Torpedo/DER, NISBD, SKIN, flb, top/flb, skin organ, l(2)57EFa, Otf11, Fam91a1, keratinized cell of epidermis, NISBD1, 9030024J15Rik, D-EGFR, l(2)57Ea, cd156b, AV220772, skin and subcutaneous tissue, NISBD2, DER1, Elp-B1RB1, Oct-11a, Tace, Elp, Wa5, l(2)09261, TGF-alpha receptor activity, ERBB, dEGFR1, DEgfr, total expressed protein, skin zone, integumental organ, CG10079, skin plus hypodermis, torpedo/egfr, skin region, Oct11, TACE, epidermal growth factor receptor activity, wa2, DmelCG10079, BC033609, Proteomes., Skin-1a, DER/top, tegument, tace, Skin, egfr, adam18, HD-33, El, top/DER, EGFR, DER/EGFR, EGfr, EgfR, dEgfr, vertebrate integument, DER flb, ERBB1, wa-2, Keratinocyte, Torpedo/Egfr, CD156b, mKIAA0493, Epoc-1, Degfr, portion of skin, vertebrate epidermis, Egf, EFG-R, integument, DEL, Errp, Erbb, DEGFR, l(2)57DEFa, l(2)05351, Der, DER, dEGFR, transforming growth factor-alpha receptor activity, entire integument, DmHD-33, dermal system, CD156B, DER/torpedo, HER1DER/faint little ball, dermis plus epidermis plus hypodermis, receptor tyrosine-protein kinase erbB-1, the integument, malpighian cell, csvp, region of skin, D-Egf, pelt, proto-oncogene c-ErbB-1, Otf-11, Skn-li, C-erb, Elp-B1, der, Elp-1, mor1, Errb1, mENA, EK2-6, epidermis, PIG61, epidermal growth factor-activated receptor activity, AI552599, c-erbB, EGFr, Egfr, ADAM18, EGF receptor activity, torpedo/Egfr, TOP, CSVP, Egf-r, d-egf-r, integumentum commune, Skn-1a, EGF-R, skin, dermoid system, entire skin, top, Torpedo/DER, NISBD, SKIN, flb, top/flb, skin organ, l(2)57EFa, Otf11, Fam91a1, keratinized cell of epidermis, NISBD1, 9030024J15Rik, D-EGFR, l(2)57Ea, cd156b, AV220772, skin and subcutaneous tissue, NISBD2, DER1, Elp-B1RB1, Oct-11a, Tace, Elp, Wa5, l(2)09261, TGF-alpha receptor activity, ERBB, dEGFR1, DEgfr, total expressed protein, skin zone, integumental organ, CG10079, skin plus hypodermis, torpedo/egfr, skin region, Oct11, TACE, epidermal growth factor receptor activity, wa2, DmelCG10079, BC033609, Proteomes., Skin-1a, DER/top, tegument, tace, Skin, egfr, adam18, HD-33, El, top/DER, EGFR, DER/EGFR, EGfr, EgfR, dEgfr, vertebrate integument, DER flb, ERBB1, wa-2, Keratinocyte, Torpedo/Egfr, CD156b, mKIAA0493, Epoc-1, Degfr, portion of skin, vertebrate epidermis, Egf, EFG-R, integument, DEL, Errp, Erbb, DEGFR, l(2)57DEFa, l(2)05351, Der, DER, dEGFR, transforming growth factor-alpha receptor activity, entire integument, DmHD-33, dermal system, CD156B, DER/torpedo, HER1liquid chromatography tandem mass spectroscopy, Formol, ANT-C, ion, Bhlha41, Scx, SCX, Lysine Hydrochloride, determination, Antp1, AUTSX5, cleavage, A4, bHLHa48, Visible Light, protein, CCN3, QM, 2-Propanone, peptide, Polypeptides, protein polypeptide chains, Lysine Acetate, Drug Tolerance, ClvPrd, peptido, diseases, scx, Hot, Methanal, hnu, B1, AntP1, Heat, 2, diseases and disorders, protein aggregate, corium, foton, fs(1)M104, WMS, L Lysine, human disease, F, LCMSMS, peptides, K, Formaldehyd, E430039A18Rik, Oxomethane, epsilon-diaminocaproic acid, proteins, outer epithelial layer, T6G21.3, number of, xscleraxis, BG:DS07700.1, IBP-9, LYS, column, ppm, Lysin, bHLHa41, sample, D1, ANT-P, s, Homo sapiens disease, stage, house mouse, Half Cystine, NOVh, GPHYSD2, drug tolerance, gamma, SGS, ratio, LC-MS2, Stars, Radiation, Zinc Cysteinate, immune system tolerance, T5E21.12, acetylation, mouse, LC-MS/MS, Light, extra or missing physical or functional parts, results, Aus, ACMICD, LIGHT, Ionen, Algorithm, BB114693, lysine, Immunologic Tolerance, NOV, PlexA1, T5E21_12, Visible Radiations, parent ion, Visible Radiation, Plxn1, HVEML, MS/MS, mice, Tails, proportionality, nov, rate, L-Cysteine, MASS, alpha, beta Trypsin, mKIAA4053, fs(1)Y[b], experimental procedures, centroiding, disease, vertebrate epidermis, biological replicate, DMDA, light quantum, Self Tolerance, Tolerance, precursor ion, liquid chromatography-tandem mass spectroscopy, C130088N23Rik, Hot Temperatures, Mouse, Polypeptide, DXS648E, Trypsin/R, STARS, hypoderm, Proteomes, centroid mass spectrum calculation, other disease, Cysteine Hydrochloride, YB, experimental, False, developmental stage, number, FBN, DMANTPE1, Gene, high weight, precursor, Antp P1, Antp P2, protein-containing complex, SCXB, presence, TYPE, epidermis, SCXA, LC-MS-MS, Buffer, Ly113, DAGA4, biological_replicate, Yb, polypeptide chain, Publication, Ms1, ECTOL1, Hu, LC-MSMS, heavy, detached, Gene Products, disease or disorder, light, Lysine, Half-Cystine, MAM, 3.4, SCG3, CG2706, Corium, ANTC, proportion, antp, iones., cutis, methods, skin, MS1, Lichtquant, MS2, iones, L Cysteine, experimental section, Formaldehyde, beta-Trypsin, Acetylations, TPP, labeling, Visible, IGFBP9, ions, CG1028, CD156, OCTD, non-neoplastic, Formalin, 10^[-6], L10, male sterility 1, has or lacks parts of type, photon, disorder, peptidos, epidermis (sensu Metazoa), TR2, AntP, ANTP, PTHB1, data, protein complex, DmelCG2706, Proteins, disorders, total expressed protein, DXS648, medical condition, hypodermis, backward, DmAntp, buffer, CD258, Peptide, LGMD2C, mereological quality, polypeptide, formaldehyde, count in organism, LC/MS/MS, Experiment, Temperatures, Immune Tolerance, native protein, natural protein, Mus, IGFBP-9, Acetate, C18, Protein, chemical analysis, condition, FORMALIN, MFS1, Clostripain, l(3)84Ba, L-Lysine, tandem MS, FORMALDEHYDE, Data Base, WMS2, Radiations, Temperature, Ant, DMDA1, distinct, MALE STERILITY 1 PROTEIN, HVEM-L, Photoradiation, Striated muscle activator of Rho-dependent signaling, Tripcellim, CD156a, Methylene oxide, sample population, LTg, Protein Gene Products, Gene Proteins, Trypure, Ns, Photoradiations, DmelCG1028, Oxomethylene, Ion, vertebrate dermis, SCARMD2, cardinality, SSKS, EG:95B7.8, quotient, liquid chromatography tandem mass spectrometry, Peptid, 2600013D04Rik, DRO15DC96Z, assay, outer epidermal layer, quantitative, 6-diaminohexanoic acid, Enisyl, Scl, reversed, Immunological Tolerance, presence or absence in organismDER/faint little ball, Networks, protein translation, Kinship, HSN1E, second cervical vertebra, determination, Laboratory, Mus domesticus, Glutaminyl-Peptide Gamma-Glutamyltransferases, Elp-B1, der, DER1., Elp-1, cleavage, A4, Errb1, CASP-14, protein, House Mouse, cell associated, 10-dimethyl-3-(1-methylethyl)-11-oxabicyclo(8.1.0)undec-6-en-2-yl ester, binding site, PIG61, axis vertebra, protein polypeptide chains, Roles, kinky, epidermal growth factor-activated receptor activity, AI552599, Concepts, Life Cycle, 3, 4, d-egf-r, protein aggregate, multicellular organismal biosynthetic process, Family Life Cycle, AI266795, increased, single-organism biosynthetic process, Kinship Network, activation inducer molecule, anabolism, Kininogenase, C2, membrane region, Swiss Mice, proteins, NISBD, flb, Protein Glutamine gamma Glutamyltransferases, vertebra 2, signaling process, Role Concepts, AI316800, keratinized cell of epidermis, Protein-Glutamine, NISBD1, 9030024J15Rik, D-EGFR, fused to, Kallikrein Light Chain, Family Life Cycles, beta Kallikrein, NISBD2, single organism signaling, DER1, protein anabolism, Ligand, protein biosynthetic process, Elp, Wa5, l(2)09261, DEgfr, mouse, integral to membrane, beta-Kallikrein, C2 vertebra, S100, S100-B, membranous organ component, CG10079, results, torpedo/egfr, Role Concept, Kallikrein B', wa2, Callicrein, Mini-ICE, Glutaminyl Peptide Gamma Glutamyltransferases, Kallidinogenase, Role, protein formation, Filiation, region, Hydrogen Oxide, Kallikrein A, egfr, adam18, Mus musculus, early T-cell activation antigen p60, ADCADN, membrane of organ, HD-33, El, positional polypeptide feature, Caspase-14 subunit p10, 5-trimethoxy-, mice, EGFR, DER/EGFR, EGfr, EgfR, Tails, FU, dEgfr, Swiss Mouse, DER flb, Caspase-14 subunit p19, gamma-Glutamyltransferases, Torpedo/Egfr, axis [C II], MICE, 4-(acetyloxy)-6, merged with, Degfr, domesticus, Life Cycles, DMDA, Kalliginogenase, protein synthesis, DEGFR, l(2)57DEFa, l(2)05351, Protein-Glutamine gamma-Glutamyltransferases, Fu, beta-Kallikrein B, dEGFR, Mouse, DmHD-33, fused, cell bound, CD69, Proteomes, accessory, Benzoic acid, FUSED, receptor tyrosine-protein kinase erbB-1, 3.4.22.-, knobbly, csvp, malpighian cell, biological signaling, D-Egf, Family Member, proto-oncogene c-ErbB-1, Transglutaminase, Filaggrin, region or site annotation, Peptidomics, Protein-glutamine:amine gamma-glutamyltransferase, alpha-Kallikrein, C-erb, mor1, Fused, EA1, Gene, mini-ICE, stalk, Gamma-Glutamyltransferases, biosynthesis, Network, mENA, MLR-3, EK2-6, protein-containing complex, terminal differentiation, supernumerary, TYPE, DAGA4, Light Chain, AIM, polypeptide chain, House, integral component of membrane, c-erbB, EGFr, Egfr, S100-alpha, Gene Products, ADAM18, Mus musculus domesticus, EGF receptor activity, torpedo/Egfr, TOP, CSVP, Egf-r, MAM, early activation antigen CD69, Mice, SCG3, study, EGF-R, positional, formation, Research, Swiss, Dilminal, ligand, axis (CII), top, Torpedo/DER, Maintenances, culm, top/flb, synthesis, S100beta, alpha Kallikrein, leukocyte surface antigen Leu-23, Enzyme, Stratum Corneum Basic Protein Precursor, Kallikrein, l(2)57EFa, NEF, region of membrane, C-type lectin domain family 2 member C, site, Kinin Forming Enzyme, l(2)57Ea, mKIAA1278, Kinship Networks, Kb, cd156b, Family, Ki, Elp-B1RB1, Filaggrin Protein, membrane, Profilaggrin, Tace, Family Research, TGF-alpha receptor activity, axis, protein complex, ERBB, dEGFR1, Proteins, DNMT, Stratum Corneum Basic Protein, total expressed protein, MCMT, B930045J24, LGMD2C, Concept, Family Members, BL-AC/P26, Axin, cervical axis, TACE, epidermal growth factor receptor activity, native protein, DmelCG10079, natural protein, Mus, Protein, chemical analysis, whole membrane, protein biosynthesis, DER/top, tace, CXXC9, transmembrane, Padutin, DMDA1, top/DER, S100a, INSDC_feature:misc_binding, increased number, cervical vertebra 2, Cathepsin, beta Kallikrein B, House Mice, ERBB1, wa-2, Keratinocyte, CD156b, signalling, Laboratory Mice, Protein Gene Products, Gene Proteins, present in greater numbers in organism, Egf, EFG-R, signalling process, binding_or_interaction_site, GP32/28, Errp, DEL, Families, S100A, Erbb, joined with, Glutaminyl-Peptide, SCARMD2, Der, DER, transforming growth factor-alpha receptor activity, assay, Relatives, 1700112N14Rik, CD156B, Laboratory Mouse, coalesced, DER/torpedo, CLEC2C, HER1, Kinin-Forming, Kinin-Forming Enzyme0trueThe Skin Barrier Defects Caused by Keratinocyte-Specific Deletion of ADAM17 or EGFR are Based on Highly Similar Proteome and Degradome AlterationsADAM17 and EGFR are essential key players for epidermal integrity. Keratinocyte-specific deletion of ADAM17 in mice results in pronounced alterations in terminal differentiation of keratinocytes leading to severe epidermal barrier defects with enhanced transepidermal water loss. Thereby, mice deficient for ADAM17 in keratinocytes phenocopy mice with a keratinocyte-specific deletion of EGFR, highlighting the role of ADAM17 as a “ligand sheddase”, as it sheds membrane bound EGFR ligands from the cell surface and finally modulates EGFR signaling. In this study we aim for the first proteomic / degradomic approach to characterize the disruption of the ADAM17-EGFR signaling axis and its consequences for epidermal barrier formation. Proteomic profiling of the epidermal proteome of mice deficient for either ADAM17 or EGFR in keratinocytes at postnatal days 3 and 10 revealed highly similar protein alterations for ADAM17 and EGFR deficiency. These include massive proteome alterations of structural and regulatory components important for barrier formation, like transglutaminases, involucrin, S100 protein family members and S100 fused-type proteins, such as filaggrin, filaggrin-2 and hornerin. Cleavage site analysis using TAILS reveals, among other ADAM17 dependent cleavage sites, increased proteolytic processing of S100 fused-type proteins, including filaggrin-2. Alterations in proteolytic processing are supported by altered protein abundance of numerous proteases upon keratinocyte-specific Adam17 or Egfr deletion, among them kallikreins, cathepsins and their inhibitors. In addition, N-terminal proteomics indicated usage of alternative translation start sites. This study highlights the essential role of proteolytic processing for maintenance of a functional epidermal barrier. Furthermore it suggests that most defects in formation of the postnatal epidermal barrier upon keratinocyte-specific ADAM17 deletion are mediated via EGFR.2016-04-202015-07-24PXD00259744544173630910090698936160488998638804333038783209285678251249668619235554NCBITaxon:10359NCBITaxon:1313884019456579559615795970448757430064187827215235443676313511076985076224308555712158916996310306215745771216979623980310022619026915746360423213562213558833474726970498030113450614647910239297601011738181459439913163910116NCBITaxon:6157569156931307674940983341182590408172408187613838039437082725631459539901197227243230746992867883937021000589NCBITaxon:245309940627912924951803430528481255417733555119350187263178723102999903469417872469453475158727597293632997966287956481882295476157546709111510422425800295486148015431744757558462823847562287360311481736231859629825489610151098239352937251264690117467349323994611676275921659554440467767373153NCBITaxon:9615260710297221280321857318431834937601911079276280863632657551031293128118365759983830797223756174920014321382426193197106592226126070526070719662788531896067029157295455817110162652827089454