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naMass SpectrometryShotgun proteomicsNot availableProteogenomicsProteomicsSweat glandsMass spectrometryhttp://www.ebi.ac.uk/pride/archive/projects/PXD010637Sweat GlandIsolation of eccrine sweat glands Skin tissues were obtained according to approved institutional review board protocol from otherwise healthy adults undergoing unrelated reconstructive surgery at the Department of Plastic and Reconstructive Surgery at the Johns Hopkins University School of Medicine. Eccrine sweat glands were dissected out from the skin dermis as previously described. Briefly, the skin was excised vertically with sharp scissors into small pieces. The cut pieces were bathed in phosphate-buffered Ringer's solution on ice. Neutral red (10 µM) was used to stain the sweat glands. After a few minutes of incubation with neutral red, the sweat glands were viewed through a Zeiss Stemi 2000 dissecting microscope. Fisherbrand fine-pointed forceps were used to dissect eccrine sweat glands from the skin. Connective tissue and fat hanging to the eccrine sweat gland were removed using Fisherbrand dissecting needles with attached holder. Images of the eccrine sweat glands were capturedusing Zeiss microscopy camera, AxioCam lCc 1, and the Axio Vision software (Release 4.8.2). RNA extraction The dissected sweat glands were placed in RNAlater (Qiagen # 76104) immediately after they were dissected out from the skin. The collection tube was centrifuged to remove RNA later solution and Trizol was added to the sample and mixed. The samples were processed in a bead-beater (FastPrep-24, MP) at a setting of 4 m/s for 20 seconds and subsequently placed on ice for five minutes. This was repeated three times. Chloroform was then added to the samples and incubated for five minutes at room temperature. The samples were then centrifuged at 8,000 x g for 5 minutes at 4°C. The aqueous phase was separated from the organic phase containing the DNA and protein. Ethanol was added to the sample for precipitation and immediately applied to RNeasy columns (RNeasy Kit, Qiagen # 74104). The manufacturer’s cleanup procedure was followed except that the RNA sample was treated with the Turbo DNA-Free kit to remove residual DNA contamination (ThermoFisher # AM1907). The RNA was stored at -80°C. The quality of RNA was measured by RNA integrity number (RIN) using Agilent 2100 Bioanalyzer. Preparation of sweat gland proteome To maximize the identification of proteins, the sweat grand proteins were extracted by two different methods - Filter-aided sample prep (FASP) approach and the guanidine hydrochloride (GuHCl) approach. For the FASP approach, sweat glands were sonicated in 50 mM TEAB/4% SDS/10 mM DTT with Halt protease inhibitor cocktail (Thermo Scientific) for 5 min followed by heating at 95°C for 5 min. After cooling, the samples were alkylated with 30 mM iodoacetamide at room temperature for 30 min followed by centrifugation at 17,000 x g for 10 min. SDS in the sample was removed by FAST method.{Wiśniewski, 2009, Universal sample preparation method for proteome analysis} Briefly, the lysate was diluted with 50 mM TEAB/8 M urea and concentrated with centricon with 30 KDa MWCO for 40 min at RT. This buffer exchange step was repeated five times. Sweat gland proteins were digested with Lys-C at room temperature for 3 h followed by further digestion with trypsin overnight. For the GuHCl approach, sweat glands were sonicated for 5 min with 35% amplitude in 8 M GuHCl, 50 mM HEPES, pH 7.0, 10 mm dithiotreitol in the presence of Halt protease inhibitor cocktail (Thermo Scientific) followed by heating at 90°C for 3 min and centrifugation at 16,000 x g for 10 min. The pellet was resuspended in 50 mM TEAB/4% SDS/10 mM DTT and subjected to further protein extraction with barocycler for 60 cycles in which each cycle consisted of 45,000 psi at 95°C for 50 sec and atmospheric pressure at 25oC for 10 sec. The proteins alkylated with 30 mM iodoacetamide at room temperature for 15 min. The proteins were digested with trypsin at the enzyme to protein ratio of 1:50 at 37oC overnight. Following enzyme digestion, the peptides were desalted using Sep-Pak C18 cartridge. The peptides prepared through FASP and GuHCl method were fractionated into 24 fractions by basic pH reversed phase liquid chromatography. Briefly, lyophilized samples were reconstituted in solvent A (10 mM triethylammonium bicarbonate, pH 8.5) and loaded onto XBridge C18, 5 μm 250 × 4.6 mm column (Waters, Milford, MA). Peptides were resolved using a gradient of 3 to 50% solvent B (10 mM triethylammonium bicarbonate in acetonitrile, pH 8.5) over 50 min collecting 96 fractions. The fractions were subsequently concatenated into 24 fractions followed by vacuum drying using SpeedVac. The dried peptides were resuspended in 15 µl 10% FA and the entire amount was injected. The peptides prepared through barocycler were fractionated by SCX StageTip as previously reported.PrideNormalized Spectral Abundance Factor - NSAFiodoacetamide derivatized residuemonohydroxylated residueRNA-sequencing and data analysis Using 1 µg of total RNA isolated from sweat glands, library preparation and RNA-seq were performed at the Deep Sequencing and Microarray Core Facility at Johns Hopkins University. RNA-seq library was constructed using the TruSeq RNA Sample Prep Kit v2 (Illumina, San Diego, CA, USA), and 200 million paired-end reads were obtained on Illumina’s Next Seq 500 platform. Quality assessment of raw fastq data generate from Illumina was carried out using FastQC v0.11.5. Raw read pairs were subjected to processing using fqtrim v0.9.5 by trimming low quality bases (Q < 20 and length < 35 bp). The human genome version GRCh38 (hg38) was used as reference with gene annotations from GENCODE. Quality processed data was then aligned using HISAT2 v2.0.1 against the human genome. Default parameters of HISAT were used in “--end-to-end” mode. HISAT alignment was then converted to BAM and sorted using samtools v1.3.1 package. Transcript assembly and quantification were carried out using StringTie v1.3.3 package with default parameters. Splice junctions from the alignment data were detected and annotated using RSeQC v2.6.4 package. Extraction of 90 bp sequence from the exon ends of novel splice junctions was carried out followed by translation into peptide sequences for database searching. GTEx pipeline was followed to align against GRCh37 (hg19) version of human genome using GENCODE annotations. Then gene expression values were derived using RNA-SeQC v1.1.8 program and compared against the median RPKM values in GTEx tissues. Data analysis Proteome Discoverer (v 2.1; Thermo Scientific) suite was used for identification and quantitation. The tandem mass spectrometry data were searched using SEQUEST search algorithms against a human RefSeq database (version 70) supplemented with frequently observed contaminants. The search parameters used were as follows: a) trypsin as a proteolytic enzyme (with up to two missed cleavages) b) peptide mass error tolerance of 10 ppm; c) fragment mass error tolerance of 0.02 Da; d) Carbamidomethylation of cysteine (+57.02146 Da) as fixed modification and oxidation of methionine (+15.99492 Da) as variable modifications. Peptides and proteins were filtered at 1% false discovery rate. Protein abundance values were calculated using normalized spectral abundance factor (NSAF). Gene ontology analysis was performed at DAVID. For the identification of novel genes expressed in protein level, transcripts assembled from the RNA-seq data were translated into three frames and a custom database was generated. The database search was conducted in the same way as the search against Reference database except for the enzyme non-specificity on the peptide N-terminal side. For the identification of peptides mapping to novel or partially novel splice junctions, splice junction sequences were extracted with RSeQC software followed by 3 frame translation of the splice junctional sequence spanning 45 nucleotide to the 5’ side and 45 nucleotides to the 3’ side. The database search was conducted in the same way as the search against Reference database. Any peptides that had identical matches in the NR human protein database were removed.ProteomicsAkhilesh PandeyLTQ Orbitrap EliteQ ExactivePARTIALDepartment of Biological Chemistry, McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USAHomo Sapiens (human)30979791 Na CH, Sharma N, Madugundu AK, Chen R, Atalar Aksit M, Rosson GD, Cutting GR, Pandey A. Integrated transcriptomic and proteomic analysis of human eccrine sweat glands identifies missing and novel proteins. Mol Cell Proteomics. 2019 10.1074/mcp.ra118.001101chanhyun.na@gmail.comBiologicalJohns Hopkins UniversityUnited StatesThe eccrine sweat gland is an exocrine gland that is involved in the secretion of sweat for control of temperature. Malfunction of the sweat glands can result in disorders such as miliaria, hyperhidrosis and bromhidrosis. Understanding the transcriptome and proteome of sweat glands is important for understanding their physiology and role in diseases. However, no systematic transcriptome or proteome analysis of sweat glands has yet been reported. Here, we isolated eccrine sweat glands from human skin by microdissection and performed RNA-seq and proteome analysis. In total, ∼138,000 transcripts and ∼6,100 proteins were identified. Comparison of the RNA-seq data of eccrine sweat glands to other human tissues revealed the closest resemblance to the cortex region of kidneys. The proteome data showed enrichment of proteins involved in secretion, reabsorption, and wound healing. Importantly, protein level identification of the calcium ion channel TRPV4 suggests the importance of eccrine sweat glands in re-epithelialization of wounds and prevention of dehydration. We also identified 2 previously missing proteins from our analysis. Using a proteogenomic approach, we identified 7 peptides from 5 novel genes, which we validated using synthetic peptides. Most of the novel proteins were from short open reading frames (sORFs) suggesting that many sORFs still remain to be annotated in the human genome. This study presents the first integrated analysis of the transcriptome and proteome of the human eccrine sweat gland and would become a valuable resource for studying sweat glands in physiology and disease.Integrated Transcriptomic and Proteomic Analysis of Human Eccrine Sweat Glands Identifies Missing and Novel Proteins.Na Chan Hyun CH, Sharma Neeraj N, Madugundu Anil K AK, Chen Ruiqiang R, Aksit Melis Atalar MA, Rosson Gedge D GD, Cutting Garry R GR, Pandey Akhilesh AMethane, 4-Dithiothreitol, Ribonucleic, DmelCG9412, Hypodermic Needles, IL1BC, CT11259, MeCN, Bhlha41, Scx, SCX, fond, CASP-1, bHLHa48, SDH, Basodexan, CycEI, Solvent, Medical Specialties, trichloro-, SDS, Polypeptides, protein polypeptide chains, KL receptor activity, hydrogen chloride, scx, Method, sense of sight, GRP1, CH3-C#N, Grp1, B1, SCO5, SEC, Urea, 1, 2, SCO1, Software Engineering, Il1bc, Gsfsow3, Non Polyadenylated, adult, integumentum commune, (R*, Multicase, thymus nucleic acid, MINT, Hypodermic, Grain, PTPSTEP, Atmospheric, sec, Tissue, Salt, entire skin, proteins, W, procedures, Hand-Held Microscopy, number of, 4-dithiothreitol, IL-1BC, SURGICAL AND MEDICAL PROCEDURES, Fast, AI047805, isolation and purification, Dehydrated ethanol, Wasserstoffchlorid, column, medicine, GP4, bHLHa41, sample, D1, Otf11, Fam91a1, Medicine, Red, CT, Bs, AV220772, skin and subcutaneous tissue, FAST, GPH, spiritus vini, Neutral, Hypodermic Needle, Caspase-1 subunit p10, l(2)gd-1, ysb, sudoriferous gland, Trypsin/K, RIBA, Ct, Liquid Chromatography, Guanidine Monohydroiodine, Light, FASTK, Procedure, Cyc E, Software Tools, DmelCG7788, Absolute Alcohol, Medical Speciality, Computer Applications, PBT, BG:DS07108.3, Dm1, monohydrochloride, Computer Applications Software, Chlorwasserstoff, vision, University, Ribonucleic acids, desoxyribose nucleic acid, l(2)05206, [OEtH], l(2)k07918, Sweats, glandula sudorifera eccrina, Acid, pbt, Software Applications, CG11628, Source Software, CHDS7, l(2)gd-l, Guanidine Phosphate, Striatum-enriched protein-tyrosine phosphatase, Methodological, study protocol, lysed material, beta Trypsin, alcohol etilico, Methylcarbinol, Hand Held Microscopy, Sox-2, Applications, Drice, Ft, ds DNA, Thrombospondin receptor, entire integument, DNA, Rin, RIN, methyl cyanide, chlorane, DRICE, rasp, PEP, 4-Dimercapto-2, Computer Software Applications, krk1, F10B6_15, rac-Dithiothreitol, region of skin, l(1)VE614, DNS, GRP1/cytohesin 1, cycline, (Deoxyribonucleotide)n, Otf-11, chlorure d'hydrogene, threo-1, DrIce, DrICE, eccrine gland, Cleland's reagent, DmcyclinE, etanol, number, STK10, Carbamide, splitted from, Antp P1, Antp P2, BDPLT10, hFat1, protein-containing complex, Dm Fat, SCXB, F10B6.15, CG7826, SCXA, CG11633, cdi7, Medical Specialities, Guanidium, CDHF7, cyclinE, Deoxyribonucleic Acid, isolation, Hu, PAS-4, Gene Products, CG7835, Cdi7, CDI7, Scarb3, CG42273, CG3352, Carmol, GP3B, 3.4, Medical, Technique, Corium, Application, Skn-1a, T6K12_14, Open Source Softwares, textus connectivus, Monohydroiodine, Toluylene Red, skin, Academic, interventionDescription, Hydrogen chloride, Tissues, Software Application, ethanol, PCE-2, beta-Trypsin, Monosodium Salt, Open Source Software, Double Stranded, 4733401P19Rik, Deoxyribonucleic acid, DmcycE, SKIN, kf, Turbo <genus>, CG1028, Solution, Computer Software Application, Study, skin organ, C2H5OH, Sep-Pak C18, drugs, Monohydrobromide, Guanidinium Chloride, Tools, Caspase-1 subunit p20, bodyfat, peptidos, operative procedures, (Deoxyribonucleotide)m, Insurance, collection tube, CPH, 4-(2-hydroxyethyl)-, PTHB1, 3-butanediol, DmelCG3352, proto-oncogene c-Kit, RNA, Guanidine Monohydrate, Guanidine Nitrate, Oct-11a, 1-(14)C-labeled, caspase 3, DNAn+1, Proteins, D10Wsu136e, BcDNA.LD22582, skin zone, intraoperative procedures, l(2)gd2, stepk, RNS, chloridohydrogen, CE-2, Needles, backward, acetonitrile, DmAntp, buffer, ME5, Dithiothreitol, Ringers Solution, Tool, polypeptide, skin plus hypodermis, glandula sudorifera merocrina, l(2)k02514, Temperatures, native protein, DmCycE, PASIV, N8, carbonyldiamide, KIT ligand receptor activity, C18, Simple Microscopy, Pressure, chemical analysis, alcool ethylique, rel-(2R, Nitrate, PAS IV, attached, Skin, DTL, XKrk1, T6K12.14, Platelet collagen receptor, Intervention, Guanidine Monohydrobromide, AU020952, Bindegewebe, DTT, Guanidine, Ant, l(2)k02602, Guanidine Sulfate, underdeveloped, Compound Microscopy, lcc, Ntup1, Chloride, SWDS, IL-1 beta-converting enzyme, Ribonucleic Acid, ethanenitrile, Interleukin-1 beta-converting enzyme, CD117, DmelCG11628, Epoc-1, ME-IV, plan specification, HIAA0929, Computer Programs, Gene Proteins, D-CycE, Ns, Applications Softwares, DmelCG1028, C-Kit, adipose, Compound, Ssm, DNA Contaminations, Desoxyribonukleinsaeure, xkl-1, l(2)fat, DRO15DC96Z, PE, phosphates, C.I. Basic Red 5, quantitative, AI450383, Sulfate, l(2)79/18, hydrochloric acid, Atmospheric Pressures, reversed, the integument, ANT-C, Grain Alcohol, pelt, NCMe, determination, PreP, Skn-li, affixed to, Antp1, CG7788, crice, Monohydrate, Guanidine Sulfate (2:1), HEPES Monosodium Salt, Ringer Solution, l(2)ft, protein, Xkl-1, Hydrogenchlorid, STEMI, Pi, AA410010, 1-hydroxyethane, alcohol, peptide, Speciality, Cut, Ethanol, Techniques, Light Microscopy, Gsfsco1, Ccne, peptido, l(1)7Ba, Specialities, l(1)7Bb, Alkohol, AntP1, Pop, Min, Cph1, protein aggregate, preoperative procedures, ST Elevation Myocardial Infarction by EKG Finding, fatty tissue, corium, Gsfsco5, fat tissue, Computer Program, RNA Gene Products, DmelCG42273, SOW3, HEPES Monosodium, Insurance Medicine, kDa, peptides, Ethyl, E927b, Simple, Open, dermoid system, hypoplasia, Computer Programs and Programming, min, cg9412, mAPC, purification, Phosphate, Aethanol, Fatty acid translocase, 3R)-1, [CH2Me(OH)], free, xscleraxis, CG11387, BG:DS07700.1, Methodological Studies, SOLO DANCERS, 1-Piperazineethanesulfonic acid, ICE, discontiguous, Hand-Held, l(2)k08110, Contamination, ANT-P, Double-Stranded DNA, Medicines, Guanidine Sulfate (1:1), TEAB, deoxyribonucleic acids, Needle, DNAn, Sl, 8-Phenazinediamine, Specialties, Monohydrochloride, BcDNA:LD22582, Alcohol, ice, iCE, drice, ribose nucleic acid, merocrine sweat gland, sample tube, ribonucleic acids, Dmel_CG7826, AI836084, Tr-kit, integumental organ, extra or missing physical or functional parts, 4-dimercapto-2, 4-disulfanylbutane-2, Double-Stranded, Ethyl alcohol, br37, CD36, Optical Microscopy, Thermo Scientific, Sweat, Source Softwares, ur, Aus, Programs, (Deoxyribonucleotide)n+m, Program, MP-1, carbamide, Gland, drIce, drICE, Ribonukleinsaeure, BB114693, BC033609, 3.1.3.48, Specialty, Computer Applications Softwares, kl1-A, pentosenucleic acids, Softwares, BcDNA:GH10590, KIT, CG9412, ETHANOL, Adults, Dmel_CG7835, Toluylene, Mnb, MNB, tyrosine-protein kinase Kit, Turbo <subgenus>, adults, EtOH, Review, Step, P45, HCl, RBM15C, Guanidine Monohydrochloride, AI047692, kit, mKIAA1104, AW124434, Methodological Study, mKIAA0493, Trichloromethane, Insurance Medicines, R*)-1, H2NC(O)NH2, vertebrate epidermis, sensory visual perception, l(2)fd, l35Dd, phosphate, phosphate ions, Faf, STEP, Acetamide, Nuclear Fast Red (Basic Dye), MP1, p45, Microscopy, Polypeptide, Review of Reported Cases, HEPES, FAT, Fat, dermal system, Proteomes, 1728, invasive procedures, dermis plus epidermis plus hypodermis, SCARB3, Procedures, SCF receptor activity, GPIIIB, DMANTPE1, Gene, cycE, fat, Computer, 3-trimethyl-, presence, l(2)br37, Deoxyribonucleic acids, epidermis, CYCLE, Intervention or Procedure, Buffer, Karbamid, cytohesin/GRP1, Aethylalkohol, method, reduced, polypeptide chain, scfr, method used in an experiment, portion of connective tissue, Harnstoff, Studies, tiny, Connective Tissues, SCFR, cloruro de hidrogeno, Guanidium Chloride, Sweat Gland, ANTC, Fdc, antp, Glands, cutis, CYCE, Plastic, Absolute, Nts, DmelCG3938, CyclE, Selb, m/s, l(2)SH2 0323, 3938, Interventional, 79/18, Connective, CG1945, Non-Polyadenylated RNA, l(2)k05007, dm-cycE, Optical, skin exudate, lysate, Neural-specific protein-tyrosine phosphatase, DmelCG11387, Roc2, ROC2, ATP:Fas-activated serine/threonine protein phosphotransferase activity, has or lacks parts of type, Glycoprotein IIIb, N-2-Hydroxyethylpiperazine-N'-2'-ethanesulfonic Acid, AntP, ANTP, sudoriparous gland, SHARP, Guanidine Hydrochloride, ST Elevation Myocardial Infarction by ECG Finding, 2-iodo-, small, Applications Software, Intervention Strategies, Open Source, DYRK1, Computer Software, bead, 3H-labeled, Medical Specialty, R74677, protein complex, total expressed protein, Scl., Guanidinium, Peptide, mereological quality, Dithiotreitol, [HCl], count in organism, Software Tool, secret agent, skin region, Oct11, Review Literature, Platelet glycoprotein IV, peroperative procedures, divided_from, natural protein, Contaminations, yeast nucleic acid, Protein, DL-threo-1, Hydrochloride, Guanidine Sulfite (1:1), Dyrk1, Skin-1a, CES2A1, techniques, CyeE, Vacuums, l(3)84Ba, ds-DNA, tegument, Software, l(2)SH0323, Cph-1, ACETONITRILE, CDHR8, cyanomethane, Ethyl Alcohol, CYH1, ribonucleic acid, c-KIT, Leukocyte differentiation antigen CD36, uree, 3-diol, operative therapy, Non Polyadenylated RNA, urea, CC1, Engineering, l(2)35Dd, vertebrate integument, Non-Polyadenylated, Tripcellim, operations, metre per second, 2310012C15Rik, N 2 Hydroxyethylpiperazine N' 2' ethanesulfonic Acid, CGI-97, sample population, UREA, incised, Interleukin-1 beta convertase, Protein Gene Products, Trypure, portion of skin, hydroxyethane, c-kit, Pressures, integument, fatty depot, glandula sudorifera, DmelCG1945, vertebrate dermis, cardinality, 3.4.22.36, perioperative procedures, l(2)24Da, Peptid, assay, GPIV, CG3938, methodology, presence or absence in organismProtein Gene Products, Sweat Gland, Gene Proteins, Glands, human being, Gland, Protein., Protein, proteomic analysis, Gene Products, Proteins, Gene, proteins, man, human, Sweatprotein translation, Ribonucleic, Bru, GRP49, BamF, Gene Ontology Projects, Materials, biml, Raw, BamC, Tandem, determination, PreP, bam, atado, Xkl-1, protein, AA410010, Mass Spectrometry Mass Spectrometry, peptide, Polypeptides, KL receptor activity, L-Isomer Methionine, protein polypeptide chains, Gsfsco1, Drug Tolerance, Methionine, ClvPrd, peptido, Pop, SCO5, Project, SCO1, rectum adenocarcinoma, Software Engineering, Analysis, DmelCG10422, protein aggregate, Gsfsow3, Gsfsco5, Non Polyadenylated, WMS, Computer Program, RNA Gene Products, SOW3, Svc, 2810441M03Rik, me75, F, Gene Expressions, peptides, Analyses, Genomes, HCAP, M, Open, Tissue, human protein, Computer Programs and Programming, nucleic acid library preparation, Mass Spectrometry, proteins, W, CG12352, CDLS3, D17Mit170, T1, hNAT5, hSAN, Human Genomes, sample, Bs, Half Cystine, 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construction, Open Source Software, Expressions, man, BMH, Computer Software Application, Non-Polyadenylated RNA, OCTD, 2-amino-4-(methylthio)butanoic acid, Tools, DmelCG12352, breakpoint, Nat5, NAT5, peptidos, Expression, FEX, CSPG6, metionina, HHT1, span, Applications Software, proto-oncogene c-Kit, Open Source, RNA, data, HG38, cou, Computer Software, DL-Methionine, protein complex, mak3, Proteins, D10Wsu136e, aligned to, RNS, BOD, L-Isomer, Cistrons, Peptide, Tool, polypeptide, read, AW112078, Software Tool, Lr, Immune Tolerance, native protein, Human Genome, natural protein, anatomical junction, KIT ligand receptor activity, yeast nucleic acid, Protein, chemical analysis, core, sequence, protein biosynthesis, MFS1, bod, Software, Library, bimel, tandem MS, Data Base, WMS2, Met, XKrk1, boundary, San, SAN, Ontology Project, ribonucleic acid, c-KIT, Col4a-1, library construction, exonic region, Non Polyadenylated RNA, Ntup1, Engineering, Non-Polyadenylated, Tripcellim, 2310012C15Rik, Ribonucleic Acid, Mass Spectrometry-Mass Spectrometry, CD117, sample population, primary structure of sequence macromolecule, Protein Gene Products, san, Computer Programs, Trypure, Gene Proteins, c-kit, Applications Softwares, C-Kit, Data, Bam-C, Ssm, BAM, SSKS, Bam, xkl-1, Bra, alpha-amino-gamma-methylmercaptobutyric acid, quotient, Peptid, PE, assay, AI450383, Mak3, ORW1, variable, Data Analyses, MAK3, Immunological ToleranceExcessive sweating, Water, Cystic fibrosis NOS, the integument, Materials, artificial sequence, pelt, determination, Skn-li, Wound, susceptibility to chronic infection by, Transient receptor potential protein 12, Meconium obstruction of intestine in mucoviscidosis, Gene Expression Profile, Profiles, protein, injury, Reading Frame, trauma, inadequate, peptide, SMAL, Polypeptides, Readability, protein polypeptide chains, halite, peptido, diseases, Roles, ionic compounds, Concepts, Heat, Msal-1, diseases and disorders, ORFs, Increased sweating, protein aggregate, Unassigned, modifier of, Prickly Heat, Unassigned Reading Frame, heat rash, integumentum commune, increased, Unidentified Reading Frames, human disease, reference sample, peptides, Genomes, Osm-9-like TRP channel 4, Open, Salt, dermoid system, entire skin, Wound Epithelialization, Bodily, Msal, proteins, VRL-2, Salz, sel, Signatures, cystic fibrosis with gastrointestinal manifestations, salts, cloruro sodico, Fibrocystic disease, salt, CF, Protein Coding, Secretion, Human Genomes, Expression Signature, Secretions, Role Concepts, Otf11, Fam91a1, Transcriptomes, Medicine, Homo sapiens disease, Pulmonary Cystic Fibrosis, Vanilloid receptor-related osmotically-activated channel, Healing, Hyperidrosis, AV220772, Protein Coding Regions, skin and subcutaneous tissue, Sweating profusely, rock salt, Water Stress, merocrine sweat gland, Prickly, external secretion, Expression Profiles, Mucoviscidosis, Cystic fibrosis NOS (disorder), RNA-seq, integumental organ, VROAC, glandula exocrina, Sweat, CF - Cystic fibrosis, exocrine gland fluid/secretion, Gene Expression, shortened, Bodily Secretion, common salt, Experimental, prickly heat, Role Concept, Gland, Expression Signatures, CYSTIC FIBROS W/O ILEUS, BC033609, Diseases, Role, Genetic Materials, secretion, Pancreatic, Fibrosis, Region, Expression Profile, END, Re Epithelialization, Genetic Material, Transcriptome Profiles, in cystic fibrosis, Sweats, glandula sudorifera eccrina, Cystic fibrosis (disorder), ORF, Pancreatic Cystic Fibrosis, Spalt, ionic compound, cystic fibrosis, natrii chloridum, human, mKIAA0493, Fibrocystic Disease of Pancreas, disease, SYNTHETIC CONSTRUCT sequences, vertebrate epidermis, Material, mucoviscidosis, Pancreas Fibrocystic Disease, artificial, Cistron, medical condition., Polypeptide, entire integument, short, exocrine gland secretion, dermal system, Proteomes, cystic fibrosis with pulmonary manifestations (disorder), dermis plus epidermis plus hypodermis, other disease, region of skin, human being, exocrine gland fluid, Healings, VRL2, exocrine gland fluid or secretion, Transcriptome Profile, Open Reading Frame, Otf-11, eccrine gland, Gene, Pancreas Fibrocystic Diseases, protein-containing complex, SPSMA, epidermis, Biomedical, Human, Unidentified Reading, Investigative, polypeptide chain, miliaria, CMT2C, Medical Research, Gene Products, Unassigned Reading Frames, disease or disorder, HMSN2C, cystic fibrosis with other manifestations (disorder), Whole Transcriptome Shotgun Sequencing, cystic fibrosis lung disease, synthetic genetic interaction (sensu inequality), sweat rash, Medical, Meconium ileus in cystic fibrosis (disorder), Pulmonary, Skn-1a, ducted gland, Sweat Gland, study, Frame, sORF, Glands, cystic fibrosis with meconium ileus, skin, Genetic, Cystic Fibrosis of Pancreas, Research, cystic fibrosis with other manifestations, Profile, Profuse sweating, Coding Region, chlorure de sodium, stubby, synthetic genetic interaction defined by inequality, cystic fibrosis with combined manifestations, Vanilloid receptor-like protein 2, man, SKIN, non-neoplastic, Unidentified Reading Frame, secreted substance, skin organ, Cystic Fibrosis, TrpV4, skin exudate, bodily secretion, sels, Kochsalz, Investigational Medicine, SSQTL1, disorder, Epithelialization, peptidos, VR-OAC, Vanilloid receptor-like channel 2, HHT1, Controlled, pseudomonas aeruginosa, Protein Coding Region, Controlling, data, Transcriptome, Oct-11a, protein complex, sales, Proteins, disorders, total expressed protein, skin zone, artificial gene, medical condition, function, synthetic DNA, Cistrons, Miliaria Rubra, Peptide, Concept, cystic fibrosis with pulmonary manifestations, polypeptide, Investigative Medicine, skin plus hypodermis, glandula sudorifera merocrina, skin region, Oct11, Temperatures, native protein, Natriumchlorid, natural protein, Human Genome, traumatic injury, Trauma, Gene Expression Signatures, chemical analysis, Protein, synthetic, condition, table salt, Gene Expression Signature, Skin-1a, Small Open Reading Frames, Diaphoresis, NaCl, tegument, Skin, Investigational, Sal, Experimental Medicine, Wound Healings, Cystic fibrosis without mention of meconium ileus, vertebrate integument, wound, B130022O04Rik, Salze, Understanding, cystic fibrosis with meconium ileus (disorder), synthetic constructs, Unidentified, sal, Epoc-1, Protein Gene Products, OTRPC4, Cystic, Gene Proteins, Sweating, portion of skin, Trp12, integument, Dietary Sodium, Gene Expression Profiles, Stress, Small Open Reading Frame, Peptid, assay, TRP12, 0610033B08Rik, Signature, ORW1, sodium chlorideExcessive sweating, Water, the integument, Materials, artificial sequence, pelt, determination, Skn-li, Wound, Transient receptor potential protein 12, Gene Expression Profile, Profiles, protein, Reading Frame, peptide, SMAL, Polypeptides, Readability, protein polypeptide chains, peptido, diseases, Roles, calcium(2+) ion, Microdissections, Concepts, Heat, diseases and disorders, ORFs, Increased sweating, protein aggregate, Unassigned, Prickly Heat, Unassigned Reading Frame, heat rash, integumentum commune, increased, Unidentified Reading Frames, human disease, reference sample, peptides, Genomes, CALCIUM ION, Osm-9-like TRP channel 4, Open, Tissue, dermoid system, entire skin, Wound Epithelialization, Bodily, proteins, VRL-2, Signatures, Protein Coding, Secretion, Human Genomes, Expression Signature, Secretions, Role Concepts, Otf11, Fam91a1, Transcriptomes, Homo sapiens disease, Vanilloid receptor-related osmotically-activated channel, Healing, Hyperidrosis, AV220772, Protein Coding Regions, skin and subcutaneous tissue, Sweating profusely, Water Stress, merocrine sweat gland, Prickly, external secretion, Expression Profiles, RNA-seq, integumental organ, VROAC, glandula exocrina, Sweat, doubly charged positive ion, exocrine gland fluid/secretion, Gene Expression, shortened, Bodily Secretion, prickly heat, Role Concept, Gland, Expression Signatures, BC033609, Diseases, Role, Genetic Materials, secretion, Region, Expression Profile, Re Epithelialization, cortex of organ, region, Genetic Material, Transcriptome Profiles, Sweats, glandula sudorifera eccrina, ORF, positional polypeptide feature, Ca(2+), human, mKIAA0493, disease, Kidneys, calcium, SYNTHETIC CONSTRUCT sequences, vertebrate epidermis, calcium(II) cation, Material, artificial, Cistron, medical condition., Polypeptide, entire integument, short, exocrine gland secretion, dermal system, Proteomes, dermis plus epidermis plus hypodermis, other disease, region of skin, human being, exocrine gland fluid, Healings, VRL2, exocrine gland fluid or secretion, Transcriptome Profile, region or site annotation, Open Reading Frame, Otf-11, eccrine gland, Gene, protein-containing complex, SPSMA, epidermis, Human, Unidentified Reading, Ca2+, polypeptide chain, miliaria, CMT2C, Gene Products, Unassigned Reading Frames, disease or disorder, HMSN2C, Whole Transcriptome Shotgun Sequencing, synthetic genetic interaction (sensu inequality), sweat rash, Skn-1a, ducted gland, Sweat Gland, Frame, study, positional, sORF, Glands, skin, Genetic, Profile, Profuse sweating, Coding Region, stubby, calcium(2+), synthetic genetic interaction defined by inequality, Vanilloid receptor-like protein 2, man, SKIN, non-neoplastic, Unidentified Reading Frame, secreted substance, skin organ, TrpV4, skin exudate, bodily secretion, SSQTL1, disorder, site, Epithelialization, peptidos, VR-OAC, Vanilloid receptor-like channel 2, Controlled, Protein Coding Region, Controlling, data, Transcriptome, Oct-11a, protein complex, Proteins, disorders, total expressed protein, skin zone, artificial gene, medical condition, function, synthetic DNA, Cistrons, Miliaria Rubra, Peptide, Concept, cortex, polypeptide, skin plus hypodermis, glandula sudorifera merocrina, skin region, Oct11, Temperatures, native protein, natural protein, Human Genome, Gene Expression Signatures, chemical analysis, Protein, sequence, synthetic, condition, Gene Expression Signature, Skin-1a, Small Open Reading Frames, Diaphoresis, tegument, Skin, Wound Healings, vertebrate integument, Understanding, synthetic constructs, primary structure of sequence macromolecule, Unidentified, Epoc-1, Protein Gene Products, OTRPC4, Gene Proteins, Sweating, portion of skin, Trp12, integument, Gene Expression Profiles, Stress, Small Open Reading Frame, Peptid, assay, TRP12, 0610033B08Rik, SignatureSweat Gland, Sweat., Glands, human being, Gland, man, human, Sweat, proteomic analysisfalseIntegrated transcriptomic and proteomic analysis of human eccrine sweat glandsThe eccrine sweat gland is an exocrine gland that is involved in the secretion of sweat for control of temperature. Malfunction of the sweat glands can result in disorders such as miliaria, hyperhidrosis and bromhidrosis. In addition, inadequate reabsorption of salt from sweat is a major feature of cystic fibrosis. Understanding the transcriptome and proteome of sweat glands is important for understanding the physiology and the role in disease. However, no systematic transcriptome or proteome analysis of sweat glands has yet been reported. To this end, we isolated eccrine sweat glands by microdissecting them from human skin and performed both RNA-seq and proteome analysis. In total, ~138,000 transcripts and ~6,100 proteins were identified. The proteome data of eccrine sweat gland showed enrichment of proteins involved in secretion, reabsorption, and wound healing while the transcriptome data did not show any enrichment for a specific pathway. Importantly, protein level identification of TRPV4 in eccrine sweat gland establishes its importance in re-epithelialization of partial-thickness wound and prevention of dehydration. Furthermore, this study enabled us to identify2 missing proteins. Integration of RNA-seq and proteomic data allowed us to identify 7 peptides from 5 novel genes. Most of the novel proteins were from short open reading frames (sORFs) suggesting that many sORFs still remain to be annotated in the human genome. The peptides mapping to the missing or novel proteins were validated by analyzing synthetic peptides. This study provides the first integrated analysis of the transcriptome and proteome of the human eccrine sweat gland and should become an invaluable resource to biomedical research community for studying sweat glands in physiology and disease.2019-04-152018-07-31PXD0106372806551646099180498511145422492156843544071284218327042363336351198343006636362758385696363121766668888111430155571145263145262364934613126957366649239161936413645106656889511363647627236611123294422295558326813042323122586365434607925193659106321432059712938655819125023254537111957748186619682158672256807617554373592536685862740816918045427877512010892250583674147956410039236942049433445974142001258641935293NCBITaxon:77643671567593643867716054048313152826930456271639162429204277944762376369612181003296292211376394863948840479NCBITaxon:215713152731051067125027820596875497792016516368407173630969893610478214836432042929410106132049138169374327732046640707453511110310780621745921612928867128867641006945694101676341021076127735113165821358430141729817463367541287356756840301350621813019411192943936711041136445913949843202530127032022149875110475941204788613474951537211097165299827905135551398031758543980126086639808137575551832019644719821114155421932555211383439119318198139818102697026635311069955529NCBITaxon:47518502983354764146547798251021691572739823897648407413735186287054824153NCBITaxon:505579844675251000364334147548021035974137081154665478983532215998381067710348198523487804225314884142228632266347946157187125939915991915729529292117527198601488406334249518691234235238152335985898598865597464998703920211107263990363527215987419082874394232418235024145113969293190259390719017463601237474636172262536001319722119158531058859698951105198997237989772000263419610264266722758858156899628737562690724420237172019998570724074767927459615333554319311671833554184005136861505968666211961748666134751536329NCBITaxon:17632862871103411033348837670580271848186536859598607575527472036105135294296110098402321000734882412336363071047005187878945556449447NCBITaxon:13356262664955419591399878168119011478281055524NCBITaxon:96063181781358584690081366848197915351930728593008526411409005118499526225412133759139014381113944235084370416744312044143708351581990119519532590841370137023943222774862615835583715371633376057267998086371137121906503714759172073127262981274575727090309031637910372814243371421044792491108239412NCBITaxon:9615658858147802692661006581373527504541356638164752037506442236695021293497905463224555253651997649908143899237453747264732376026473128870559553614169152527403271602938511497864794363755293824794372251177529912044741673872175091938441376593934347393271591002263767423444234546700128768959119846704146479293652803428038280351080348330779179392112621229502711311803738681911265269971028622527391076015973611432425071140981087135547743658142450722953379698204232124223321803369822130080347081645733185889716546759928777020382931412529550811481744888643745264999728303571830855071762997076789268370415156288765138856392456115293202086595551655181135491432137159749535026143213858781707683194963401614280053164686275110466038151845474485009747697477118051240174841855601555479204274381513201488711053253297261005941289093164351211604990013894382835423242331082934122058560416299131122731701879915429131267818187104931640729946714423683033373304852646383011306413992530642369923121162065261610337286129940207340115357652611114716133865431201725323712552329935316317NCBITaxon:11084993841063267872862321933486208621160791392227681196125522814953932132232334774121827532917573129948861814084341033519963139248913924881153381153393304393755699286265599339112094886241571839975864442374864353324442606653324134390976821271786371979856863999839986865535291486541042986357358257632884019864982380111911028688999586652203411508599632138119380625631886571303443112130886581144889198466645463NCBITaxon:11072NCBITaxon:11071142800628835374375116597145943116598854331116781077854314408043933434NCBITaxon:11082693977381382189931824431935128387145953116885311213278695berry8694100333586972443664701501114286921504751499105484204714722354025120847014649744497431170089306218644193061107632330805660396016217974825853946055396513212790568813377512981766378876017112147861050236694657349632051124501897441739412450128853183889198679488531911137910371647351660152943631631198573896939343237956026087121346334323223544314275241433611334565560291699636436802790105603140818838185733703401881274414936530437552196969716203871398558408172405598609183827275921274423408170275981274420127442623545513656531762793821176275943286303902474801199150956481882291690255120455148167238473848190695148498101510693731263753405203860186497405224052141856451804NCBITaxon:8134780091522754847938703338493871917226175610946161435120325812055052352594473205493864112572265121651253880376686120678112202230743220118670511867057387334869121938791868023980071868012154514683514081861508471206777691838851608611388869152167786721279010087043512317196928123929486771331548454226126219813188268241122926571632957693812471246148545132838834839294911390363120416266589077151488003694269416943694518790101508081705394206542105568731744712701532924281411194669116960145458695320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