Prideapplication/xmlftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/MS_identifications.xlsxftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S44.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S23.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S7.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S20.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S25.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S72.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S34.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S51.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S28.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S13.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S61.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S36.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S65.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S27.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S6.msfftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S20.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S36.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S23.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S7.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S27.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S44.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S28.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S61.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S65.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S6.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S13.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S25.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S34.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S51.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/S72.rawftp://ftp.pride.ebi.ac.uk/pride/data/archive/2017/07/PXD005780/matched_metagenomic_database.fastaprimaryOK200004860uzzau@portocontericerche.itAlessandro TancaMass SpectrometryShotgun proteomicsNot availableMetaproteomicsGut metagenomeGut metaproteomehttp://www.ebi.ac.uk/pride/archive/projects/PXD005780FecesSamples. Stool samples were collected from 15 healthy Sardinian volunteers selected to avoid age, sex and BMI biases (SI Table S1) and immediately stored at -80°C. Two equal portions were collected from each sample and subjected to DNA and protein extraction, respectively. Metaproteome analysis. Proteins were extracted (1) and digested (2) as described previously. LC-MS/MS analysis was carried out using an LTQ-Orbitrap Velos mass spectrometer interfaced with an UltiMate 3000 RSLCnano LC system (both from Thermo Scientific). (1) Tanca A, et al. (2014) A straightforward and efficient analytical pipeline for metaproteome characterization. Microbiome 2(1):49. (2) Tanca A, Biosa G, Pagnozzi D, Addis MF, & Uzzau S (2013) Comparison of detergent-based sample preparation workflows for LTQ-Orbitrap analysis of the Escherichia coli proteome. Proteomics 13(17):2597-2607.PrideSpectrum countingiodoacetamide derivatized residuePeptide identification was performed using Sequest-HT and Percolator, embedded in Proteome Discoverer™, as described previously (3). (3) Tanca A, Palomba A, Pisanu S, Addis MF, & Uzzau S (2015) Enrichment or depletion? The impact of stool pretreatment on metaproteomic characterization of the human gut microbiota. Proteomics 15(20):3474-3485.ProteomicsSergio UzzauLTQ Orbitrap VelosPARTIALPorto Conte Ricerche, Alghero, Sardinia, ItalyHuman Gut Metagenome28709472 Tanca A, Abbondio M, Palomba A, Fraumene C, Manghina V, Cucca F, Fiorillo E, Uzzau S. Potential and active functions in the gut microbiota of a healthy human cohort. Microbiome. 2017 Jul 14;5(1):79 10.1186/s40168-017-0293-3tanca@portocontericerche.itDeep learningMetaproteomicsMachine learningBenchmarkingBiologicalProteomics Laboratory, Porto Conte Ricerche, Alghero, ItalyItaly<h4>Background</h4>The study of the gut microbiota (GM) is rapidly moving towards its functional characterization by means of shotgun meta-omics. In this context, there is still no consensus on which microbial functions are consistently and constitutively expressed in the human gut in physiological conditions. Here, we selected a cohort of 15 healthy subjects from a native and highly monitored Sardinian population and analyzed their GMs using shotgun metaproteomics, with the aim of investigating GM functions actually expressed in a healthy human population. In addition, shotgun metagenomics was employed to reveal GM functional potential and to compare metagenome and metaproteome profiles in a combined taxonomic and functional fashion.<h4>Results</h4>Metagenomic and metaproteomic data concerning the taxonomic structure of the GM under study were globally comparable. On the contrary, a considerable divergence between genetic potential and functional activity of the human healthy GM was observed, with the metaproteome displaying a higher plasticity, compared to the lower inter-individual variability of metagenome profiles. The taxon-specific contribution to functional activities and metabolic tasks was also examined, giving insights into the peculiar role of several GM members in carbohydrate metabolism (including polysaccharide degradation, glycan transport, glycolysis, and short-chain fatty acid production). Noteworthy, Firmicutes-driven butyrogenesis (mainly due to Faecalibacterium spp.) was shown to be the metabolic activity with the highest expression rate and the lowest inter-individual variability in the study cohort, in line with the previously reported importance of the biosynthesis of this microbial product for the gut homeostasis.<h4>Conclusions</h4>Our results provide detailed and taxon-specific information regarding functions and pathways actively working in a healthy GM. The reported discrepancy between expressed functions and functional potential suggests that caution should be used before drawing functional conclusions from metagenomic data, further supporting metaproteomics as a fundamental approach to characterize the human GM metabolic functions and activities.Potential and active functions in the gut microbiota of a healthy human cohort.Tanca Alessandro A, Abbondio Marcello M, Palomba Antonio A, Fraumene Cristina C, Manghina Valeria V, Cucca Francesco F, Fiorillo Edoardo E, Uzzau Sergio Sliquid chromatography tandem mass spectroscopy, portionem cacas, stool, DNS, (Deoxyribonucleotide)n, determination, Untrained, Gene, bacterium E3, protein, protein-containing complex, Feces, body system, Deoxyribonucleic acids, LC-MS-MS, Volunteer Workers, protein polypeptide chains, dung, Deoxyribonucleic Acid, polypeptide chain, Detergents, Escherchia coli, LC-MSMS, Gene Products, system, Enteroinvasive Escherichia coli, Enterococcus coli, ordure, protein aggregate, portion of fecal material, portion of feces, thymus nucleic acid, anatomical systems, Sex, LCMSMS, Genotypic Sex, Escherichia/Shigella coli, Personnel, Volunteer Worker, Untrained Personnel, Double Stranded, E coli, Deoxyribonucleic acid, proteins, Worker, Phenotypic, E. coli, BMI, Eschericia coli, Enteroaggregative Escherichia coli, Table, piece of shit, sex, Voluntary Worker, sample, Voluntary Workers, Double-Stranded DNA, (Deoxyribonucleotide)m, deoxyribonucleic acids, DNAn, Volunteer Personnel, portion of dung, LC-MS2, Alkalescens-Dispar Group, Peptidomics., Bacterium coli, protein complex, DNAn+1, Proteins, fewmet, LC-MS/MS, detergent, Volunteerism, faeces, total expressed protein, cow dung, Double-Stranded, EAggEC, Enteroaggregative E. coli, spoor, Thermo Scientific, cow pat, (Deoxyribonucleotide)n+m, LC/MS/MS, native protein, natural protein, Diffusely Adherent E. coli, Bacillus coli, Voluntary, Quetelet's Index, Protein, chemical analysis, excreta, biological sex, scat, ds-DNA, connected anatomical system, desoxyribose nucleic acid, fecal material, excrement, Genotypic, Diffusely Adherent Escherichia coli, Volunteer, portion of excrement, portion of fecal matter, Enteroinvasive E. coli, portion of faeces, portion of guano, guano, spraint, sample population, Protein Gene Products, organ system, Gene Proteins, frass, Bacterium coli commune, Phenotypic Sex, liquid chromatography-tandem mass spectroscopy, ds DNA, Desoxyribonukleinsaeure, liquid chromatography tandem mass spectrometry, assay, DNA, fecal matter, Proteomes, portion of scat, droppingsdigestive canal, Microbial Community Composition, AI747421, Community Composition, gut, Microbiomes, human being, Microbial Community Compositions, lower gastrointestinal tract, Microbial Community, Microbial Community Structure, Gus-u, alimentary tract, Community Structure, Gus-t, Gus-s, enteric tract, Gus-r, Microbial, man, Human, Composition, gut tube, Gus, Gur, Gut, g, gastrointestinal system, lower GI tract, Microbial Community Structures, digestive tube, Microbiotas, asd, alimentary system, human., alimentary canal, Human Microbiomes, Microbial Communities, Community, Human Microbiome, MicrobiomePeptidomics., excrement, portionem cacas, stool, portion of excrement, peptides, portion of fecal matter, human gut microbiota, portion of faeces, portion of guano, fewmet, total expressed protein, faeces, cow dung, guano, Ximpact, Feces, spraint, spoor, Peptide, impact-a, cow pat, polypeptide, peptide, Polypeptides, dung, piece of shit, frass, ClvPrd, peptido, Peptid, peptidos, excreta, Polypeptide, scat, ordure, E430016J11Rik, fecal matter, portion of fecal material, Proteomes, RWDD5, portion of scat, portion of dung, droppings, portion of feces, fecal materialActivity, Ristella, Laboratory, Embden-Meyerhof, Ristella <CFB group bacteria>, Glykan, polysaccharide degradation, Embden Meyerhof Parnas Pathway, School-Age, solute:solute exchange, polisacarido, IMP1, diseases, Roles, T18E12.21, Concepts, diseases and disorders, Embden-Meyerhof Pathway, Research Activity, Laboratory Research, multicellular organismal biosynthetic process, Priorities, average, single-organism biosynthetic process, human disease, T18E12_21, anabolism, Microbial Community Structure, alimentary tract, Community Genomics, Glycane, Pathways, inhibition of homeostatic process, genetic, g, H13, Role Concepts, Glycan, Microbial Community Structures, homeostasis, digestive tube, asd, Homo sapiens disease, variability of a physical quality, Research Priority, alimentary system, ratio, close to, Genomics, Pathway, lower gastrointestinal tract, PSL3, SPPL1, IMPAS-1, familial, anaerobic glycolysis, Research Priorities, Normalcy, Microbial, Embden-Meyerhof-Parnas, results, Glykane, Embden-Meyerhof-Parnas Pathway, low GC Gram+, Role Concept, positive regulation of homeostatic process, single-organism transport, multicellular organismal polysaccharide catabolic process, School Age, Capsularis, Diseases, Role, Individual, Microbial Communities, Research and Development, Populations, AI747421, low G+C Gram-positive bacteria, carbohydrate metabolism, Microbial Community Compositions, Firmicutes, proportionality, rate, Low G+C firmicutes, human, Embden-Meyerhof pathway, Activities, polysaccharides, Composition, disease, Health, glycolysis, gastrointestinal system, MSTP086, small molecule transport, polysaccharide breakdown, medical condition., inherited genetic, School-Age Population, IMPAS, Population Genomics, Embden-Meyerhof Pathways, Human Microbiomes, Community, polysaccharide catabolism, digestive canal, Microbial Community Composition, other disease, human being, bioformation, conformation, clostridial firmicutes, glycans, enteric tract, biosynthesis, Normalcies, firmicutes, Human, Gus, Gur, Gut, Metabolism, single-organism carbohydrate metabolic process, disease or disorder, ATSPP, Polysaccharide, Normalities, School-Age Populations, study, proportion, DmelCG6525, multicellular organismal carbohydrate metabolic process, gut, Research, formation, Bacillus/Clostridium group, Population, man, FBgn0082831, synthesis, non-neoplastic, PSENL3, lower GI tract, negative regulation of homeostatic process, dJ324O17.1, disorder, Development and Research, School Age Population, constitutitional genetic, relational structural quality, Firmicuteota, modifed Embden-Meyerhof pathway, Human Microbiome, Autoregulation, Individual Health, Community Composition, Embden Meyerhof Pathway, signal peptide peptidase, "Firmicutes" corrig. Gibbons and Murray 1978 (Approved Lists 1980), Environmental Genomics, disorders, SPP, Spp, Concept, near to, Environmental, gut tube, Priority, Research Activities, spp, condition, regulation of homeostatic process, alimentary canal, Microbiome, polisacaridos, Microbiomes, Normality, Microbial Community, Gus-u, Community Structure, Gus-t, Gus-s, Gus-r, Bacillota corrig. Gibbons and Murray 2021, Carbohydrate, Embden-Meyerhof-Parnas pathway, School Age Populations, Bacillaeota, activation of homeostatic process, Clostridium group firmicutes, approaches, vicinity of, CG6525, quotient, Microbiotas, variable, hereditary, General activitydigestive canal, human being, bioformation, Activity, HSN1E, conformation, clostridial firmicutes, glycans, enteric tract, EA1, Embden-Meyerhof, Glykan, biosynthesis, Development, MLR-3, polysaccharide degradation, Embden Meyerhof Parnas Pathway, firmicutes, School-Age, solute:solute exchange, Gus, polisacarido, Gur, AIM, Gut, IMP1, Roles, Metabolism, Consensus, single-organism carbohydrate metabolic process, Concepts, T18E12.21, Embden-Meyerhof Pathway, early activation antigen CD69, ATSPP, Polysaccharide, multicellular organismal biosynthetic process, study, School-Age Populations, proportion, single-organism biosynthetic process, multicellular organismal carbohydrate metabolic process, DmelCG6525, gut, T18E12_21, formation, activation inducer molecule, anabolism, alimentary tract, Bacillus/Clostridium group, Community Genomics, Glycane, Pathways, Population, inhibition of homeostatic process, man, FBgn0082831, genetic, synthesis, leukocyte surface antigen Leu-23, PSENL3, g, lower GI tract, H13, Role Concepts, Glycan, C-type lectin domain family 2 member C, negative regulation of homeostatic process, dJ324O17.1, homeostasis, digestive tube, asd, Consensus Development, variability of a physical quality, School Age Population, alimentary system, constitutitional genetic, relational structural quality, Firmicuteota, modifed Embden-Meyerhof pathway, ratio, Autoregulation, data, Genomics, Pathway, Embden Meyerhof Pathway, signal peptide peptidase, lower gastrointestinal tract, "Firmicutes" corrig. Gibbons and Murray 1978 (Approved Lists 1980), Environmental Genomics, PSL3, SPPL1, IMPAS-1, DNMT, familial, anaerobic glycolysis, SPP, Spp, MCMT, Embden-Meyerhof-Parnas, results, Concept, Glykane, BL-AC/P26, Environmental, Embden-Meyerhof-Parnas Pathway, gut tube, low GC Gram+, Role Concept, positive regulation of homeostatic process, single-organism transport, multicellular organismal polysaccharide catabolic process, School Age, Drawings, Role, spp, background, regulation of homeostatic process, human., alimentary canal, CXXC9, early T-cell activation antigen p60, Metagenomes, Populations, ADCADN, polisacaridos, AI747421, carbohydrate metabolism, low G+C Gram-positive bacteria, Firmicutes, proportionality, Gus-u, Gus-t, Gus-s, Gus-r, rate, Low G+C firmicutes, Bacillota corrig. Gibbons and Murray 2021, Carbohydrate, Embden-Meyerhof-Parnas pathway, School Age Populations, Bacillaeota, human, Embden-Meyerhof pathway, introduction, polysaccharides, activation of homeostatic process, glycolysis, Clostridium group firmicutes, GP32/28, gastrointestinal system, MSTP086, CG6525, quotient, small molecule transport, polysaccharide breakdown, inherited genetic, School-Age Population, IMPAS, Population Genomics, hereditary, CD69, General activity, Embden-Meyerhof Pathways, CLEC2C, Community, polysaccharide catabolismhuman gut microbiota.486trueRelating human gut metagenome and metaproteomeGut microbiome research is rapidly moving towards the functional characterization of the microbiota by means of shotgun meta-omics. Here, we selected a cohort of healthy subjects from an indigenous and monitored Sardinian population to analyze their gut microbiota using both shotgun metagenomics and shotgun metaproteomics. We found a considerable divergence between genetic potential and functional activity of the human healthy gut microbiota, in spite of a quite comparable taxonomic structure revealed by the two approaches. Investigation of inter-individual variability of taxonomic features revealed Bacteroides and Akkermansia as remarkably conserved and variable in abundance within the population, respectively. Firmicutes-driven butyrogenesis (mainly due to Faecalibacterium spp.) was shown to be the functional activity with the higher expression rate and the lower inter-individual variability in the study cohort, highlighting the key importance of the biosynthesis of this microbial by-product for the gut homeostasis. The taxon-specific contribution to functional activities and metabolic tasks was also examined, giving insights into the peculiar role of several gut microbiota members in carbohydrate metabolism (including polysaccharide degradation, glycan transport, glycolysis and short-chain fatty acid production). In conclusion, our results provide useful indications regarding the main functions actively exerted by the gut microbiota members of a healthy human cohort, and support metaproteomics as a valuable approach to investigate the functional role of the gut microbiota in health and disease.2017-07-192017-01-25PXD005780280655164609918049829535851114513335244224942024527872842188090368093270429330845811363336351524260119834300662758385NCBITaxon:12806888856189611143011872635555711452623649346136403195206512695736664923916197467073853883641135461582158231066568895390531136364762721123294422291180992323122586365458333460792519365914320597129386558195851149315242592637371250232NCBITaxon:12721119577481866148858431043002451358672256807617554373592540816918045426683429531945305861266835149822505814963674452158554528113208147956410039237514636942049433208964208963445974137979138712728384400667203275453293529367156759388036478434643867716052165994048321659721659545501315282693042165915627161696833916243218321729204277944762376454317203094980197525556978159509439843948840479NCBITaxon:2157525338551531051067439899037122857759515205968762262311210884555588845587077918178081736309431796758144571698936160488409859NCBITaxon:371621047826794015059071483643204210613204913816936303555432046632633311994565453511025411103217459216129264201288676410026420369456941016763410210761277351524867131658239254727933428146135843014172987609174633457744491675476048932128735675684030135062191726518130194111929439367110411310239320255598632582788396591047594120347495109716049016529983552566366817585439801102432608663102459808757417555183201964471452680457966313670190832331156114155355104219325552113834397634981812359871069955529NCBITaxon:4751850298335476113615741465477180066556484337677144137929976798251021699823897641400820840741400821137351155711089725482444474153NCBITaxon:50557899698446752510003646534933414714749354801946751299114359741276473137086326547876565893639835322159589366983819201110677577271034819852348780265554479464098609958615718717668599191572951372211711012929210768783497419860202809392499148840633424923423515233598598865591324974649987039202111072943311711393635272159874190850371170827117082841823502414511396929319021978224746360461471237472269402375939723287581722532566481219071905079452659600131972211915853101774208588596NCBITaxon:615750521028307989515586411051989972375059576131720002634196102641088878176179212190659407121906772275885815689964975151950582873757244NCBITaxon:22023717NCBITaxon:170361220199985705060506172407476792745062724915961533355433355418400524643213686150596487844612286662941786661347515363291018194301086731018510184NCBITaxon:1763573039635586331492862876705801321650852718481865368595105135553482260710185687294296110091416333NCBITaxon:619184023210007348824123363630710470058127187878645159812894555663577553470951451225786425011449447260704NCBITaxon:13356262607052607076584574054163677483167233150561007133705654195NCBITaxon:305550445791399878168119011928752829126806315327341478281055524NCBITaxon:960613585846900811116781502283051930283672859300852900561742NCBITaxon:569382688306401794699151011849952622543525875913151590144824283811139442258441604193704759272120441437086042328167728503515819901195195326738637013702492405360883858162248839432186156227748817562615835583715333760572671193501998086119350216787910857913711371272407190650371475917262981288057125627457572709030903113335503728142433747924910078711082NCBITaxon:96157243359463318586260799141262200298147802492670NCBITaxon:12637192812492125283038568385695911692661006581373523756127504542666255151978813566383750644223360094129349790546322455525363524721401257515115199764990851515143899237453747445446415243235937605920222887051580595536555088157932716029385157811497863389229382894535936479437509173NCBITaxon:6854412754225117752991590109013416738715895521012966103061296715851584515849376593934327159100226376733889147923590461423451287689591198332966467041464792936576164280341160221641331599280382803536909108917688149894621812141080348179392287889295027113133295282829445686174934681911265269972273218903821076012471904007271597369978281143114020641198108713554774365814245072295337969820423212422312916321803347087145372091645733215358157546266940401379928777020206032811431933829355083329129221148643745725264999728303573908718308726727103790855071762997076789268370415573729214053651388294128160783956392431426265489822737756115596207322086595882735551693605330797255181135491432137159749535026115955614321384060725878141447335215094933491949634016143347280059360463164610466076290338151845474485003567097477265669771951240174841855601335755547920427415805360769946881037620148873297261005941200375828909316435428268314256850071230383990013894649408382835941442423221978542337742171598921409212205853939621157781154756929793244704749906604162103591905158114452749907312987779913112273170187546991103589915NCBITaxon:372961036098334187104911307981336795316407299467103401578658264530333733063830113064139925306423636505911696547885469811211620343176526161033728612994044586207340115357111952965261174553111471613386541313166343055213231201798360253237312761255232993512993850573163176426197940115134584457726787286232193341513461862086211607913922279835168119612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