panorama0000ProteomicsAndreas HoberHomo Sapienshttps://panoramaweb.org/human_secretome.urlandreas.hober@scilifelab.seScience for Life Laboratory, KTH - Royal Institute of TechnologyPanoramaPublicThe proteins secreted by human tissues and blood cells, the secretome, are important both for the basic understanding of human biology and for identification of potential targets for future diagnosis and therapy. Here, a high-throughput mammalian cell factory is presented that was established to create a resource of recombinant full-length proteins covering the majority of those annotated as 'secreted' in humans. The full-length DNA sequences of each of the predicted secreted proteins were generated by gene synthesis, the constructs were transfected into Chinese hamster ovary (CHO) cells and the recombinant proteins were produced, purified and analyzed. Almost 1,300 proteins were successfully generated and proteins predicted to be secreted into the blood were produced with a success rate of 65%, while the success rates for the other categories of secreted proteins were somewhat lower giving an overall one-pass success rate of ca. 58%. The proteins were used to generate targeted proteomics assays and several of the proteins were shown to be active in a phenotypic assay involving pancreatic β-cell dedifferentiation. Many of the proteins that failed during production in CHO cells could be rescued in human embryonic kidney (HEK 293) cells suggesting that a cell factory of human origin can be an attractive alternative for production in mammalian cells. In conclusion, a high-throughput protein production and purification system has been successfully established to create a unique resource of the human secretome.High throughput generation of a resource of the human secretome in mammalian cells.Tegel Hanna H, Dannemeyer Melanie M, Kanje Sara S, Sivertsson Åsa Å, Berling Anna A, Svensson Anne-Sophie AS, Hober Andreas A, Enstedt Henric H, Volk Anna-Luisa AL, Lundqvist Magnus M, Moradi Mona M, Afshari Delaram D, Ekblad Siri S, Xu LanLan L, Westin Malin M, Bidad Faranak F, Schiavone Lovisa Holmberg LH, Davies Rick R, Mayr Lorenz M LM, Knight Sinead S, Göpel Sven O SO, Voldborg Björn G BG, Edfors Fredrik F, Forsström Björn B, von Feilitzen Kalle K, Zwahlen Martin M, Rockberg Johan J, Takanen Jenny Ottosson JO, Uhlén Mathias M, Hober Sophia S293 cell, host organism, d230, Antemortem Diagnosis, human being, single-organism developmental process, determination, Bodies, Peptidomics, supply, postnatal development, Human Figures, Gene, dTAFII250, growth and development, Diagnosis, EfW1, Human, dmTAF[[II]]230, Readability, Homo sapiens, Secretome, dmTAF1, Taf230, Gene Products, Mass, symptoms, Screening, Projections and Predictions, HEK 293, Human Embryonic Kidney 293, Antemortem, CHO Cell, Man, TAF250, Dedifferentiations, treatment, Taf200, dTAF[[II]]250, Man (Taxonomy), TFIID TAF250, cel, distribution, cell, Tissue, Parts, Taf1p, Futurology, Diagnoses and Examinations, Body Parts and Fluids, dTAF250, Cell Dedifferentiations, disease management, Figure, Therapies, Human Bodies, Body Parts, TAF, Diagnose, Predictions and Projections, 293 HEK, Therapy, Antemortem Diagnoses, screening, dTAF[[II]]230, TAF[[II]]250, Secretomes, findings, CHO, Modern, Proteins, TAF200, l(3)84Ab, BG:DS00004.13, TAFII-250, TAF250/230, Examination and Diagnoses, Cell, predicted, Diagnoses, dTAF230, development, TAFII250, Postmortem, Screenings, Mass Screenings, p230, Examinations and Diagnoses, Protein, chemical analysis, TAF[[II]]250/230, TFIID, supply and distribution, Postmortem Diagnosis, Diagnoses and Examination, Taf[[II]]250, Postmortem Diagnoses, TAF[[II]]230, Protein., 293, postnatal growth, HEK293, Protein Secretomes, signs, Human Figure, TAF[II]250, Understanding, HEK-293, CG17603, TAF[[II]], Treatments, human, Body, Protein Gene Products, Gene Proteins, DmelCG17603, Therapeutic, Taf250, SR3-5, Figures, Modern Man, Cells, Protein Secretome, Treatment, assay, Dedifferentiation, Future, growth, TAF230, TAF1Human, Secretomes, human being, Man (Taxonomy), Homo sapiens, Secretome, Modern Man, Protein, Modern, Protein Secretome, Protein Secretomes, Cell., Man, humanHuman, Secretomes, human being, Man (Taxonomy), Homo sapiens, Secretome, Modern Man, Protein, Modern, Protein Secretome, Protein Secretomes, Cell., Man, human293 cell, ShakB, Antemortem Diagnosis, Materials, determination, Recombinant DNA, Blood, CG1321, CG32508, archinephron, protein, Diagnosis, gonada of female organism genitalia, dmTAF[[II]]230, Readability, Armenian, pass, Secretome, symptoms, 2, 3, HEK 293, 6, Human Embryonic Kidney 293, protein aggregate, CHO Cell, multicellular organismal biosynthetic process, genitalia of female organism gonada, Dedifferentiations, treatment, DNA Recombinant, single-organism biosynthetic process, thymus nucleic acid, head kidney, Man (Taxonomy), Shak B, TFIID TAF250, cel, ovum-producing ovary, Tissue, female organism reproductive system gonad, Futurology, purification, l(1)TH73, CG34358, isolation and purification, Recombinant DNA Proteins, reproductive system of female organism gonada, scientific observation, Hamsters, Chinese Hamsters, disease management, Therapies, l(1)LB21, Double-Stranded DNA, Chinese hamsters, R-9-29, deoxyribonucleic acids, DNAn, shkB, l(1)W3b, Diagnose, Predictions and Projections, 293 HEK, Therapy, screening, Pas, dTAF[[II]]230, l(1)W3, uvrc homolog protein, CHO, DNA Proteins, shB, gonada of reproductive system of female organism, gonad of female organism reproductive system, Chinese Hamster Ovary cell, Modern, TAF200, inx8, Double-Stranded, TAFII-250, TAF250/230, predicted, Diagnoses, (Deoxyribonucleotide)n+m, nj-156, pas, TAFII250, anterior kidney, Postmortem, Screenings, Ovaries, cho, Armenian Hamster, Examinations and Diagnoses, Corpuscles, Genetic Materials, Postmortem Diagnosis, Cricetulus aureus, desoxyribose nucleic acid, female reproductive system gonad, Genetic Material, W3, Diagnoses and Examination, Blood Cell, Postmortem Diagnoses, gonada of genitalia of female organism, Recombinant Proteins, 293, l(1)R-10-3, gonad of reproductive system of female organism, Protein Secretomes, signs, INSDC_feature:gene, CG17603, TAF[[II]], Treatments, l(1)R-10-7, human, EMBRYO DEFECTIVE 133, Shak-B, Grey, pronephric nephron, Corpuscle, female organism genitalia gonada, DNA Recombinant Proteins, Dmel_CG12678, Taf250, Material, pronephron, SR3-5, Cells, ds DNA, Cistron, Blood Corpuscles, DNA, Dedifferentiation, pronephric kidney, Cricetus griseus, TAF230, humans, d230, human being, DNS, (Deoxyribonucleotide)n, Peptidomics, shak-B, Gene, dTAFII250, protein-containing complex, EfW1, Deoxyribonucleic acids, Human, gonad of genitalia of female organism, TITAN 1, Homo sapiens, Deoxyribonucleic Acid, dmTAF1, isolation, Taf230, Cricetulus barabensis griseus, Gene Products, Mass, Screening, Grey Hamster, Recombinant Protein, Projections and Predictions, R9-29, Chinese hamster, TFC D, Antemortem, Blood Corpuscle, Man, Biosynthetic, Hamster, TAF250, Taf200, gonada of female organism reproductive system, anatomical systems, dTAF[[II]]250, Genetic, cell, genitalia of female organism gonad, Double Stranded, Taf1p, Deoxyribonucleic acid, Diagnoses and Examinations, Biosynthetic Proteins, dTAF250, l(1)R-9-29, Cell Dedifferentiations, reproductive system of female organism gonad, nj156, Armenian Hamsters, Dmel_CG32508, EMB133, ovary - animal, ovary, (Deoxyribonucleotide)m, DmelCG34358, TAF, Chinese, female organism genitalia gonad, female reproductive system gonada, CG15451, Grey Hamsters, Antemortem Diagnoses, measuring, TAF[[II]]250, Secretomes, findings, gonad of female reproductive system, whole blood, endonuclease cho, protein complex, synthesize, DNAn+1, gonada of female reproductive system, Proteins, l(3)84Ab, BG:DS00004.13, Cistrons, Chinese Hamster, Cell, Examination and Diagnoses, CHAMPIGNON, dTAF230, embryonic kidney, TUBULIN FOLDING COFACTOR D, native protein, Mass Screenings, p230, Protein, chemical analysis, TAF[[II]]250/230, TFIID, ds-DNA, Taf[[II]]250, CG12678, ovaries, female organism reproductive system gonada, animal ovary, TAF[[II]]230, Protein., female gonad, ovarium, HEK293, gonad of female organism genitalia, TAF[II]250, l(1)19Eb, Understanding, HEK-293, Recombinant, Protein Gene Products, Gene Proteins, DmelCG17603, Therapeutic, Modern Man, Desoxyribonukleinsaeure, Protein Secretome, l(1)R-10-14, Treatment, assay, Future, TAF1, Biosynthetic Protein, E810falseHigh throughput generation of a resource of the human secretome in mammalian cellsThe proteins secreted by human tissues (the secretome) are important for the basic understanding of human biology, but also for identification of potential targets for future diagnosis and therapy. Here, we present an annotation of all predicted secreted proteins (n=2,623) with information about their spatial distribution in the human body. A high-throughput mammalian cell factory was established to create a resource of recombinant full-length proteins. This resource was used for phenotypic assays involving β-cell dedifferentiation and for development of targeted proteomics assays. A comparison between host cells, including omics analysis, shows that many of the proteins that failed to be generated in CHO cells could be rescued in human HEK 293 cells. In conclusion, the human secretome has been mapped and characterized to facilitate further exploration of the human secretome.Mon Mar 15 00:00:00 GMT 2021PXD012054960632502629