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To pinpoint mechanisms underlying glomerulosclerosis, the authors performed ultrasensitive proteomics of purified podocyte fractions at early injury stages in mouse models of glomerular disease induced by doxorubicin or LPS. These analyses revealed an early stress response that involves upregulation of metabolic, proteostatic, and mechanoresponsive mechanisms. They also identified conserved upregulated proteins involved in the podocyte stress response, including the mechanosensor Filamin-B, and found a high correlation between proteinuria and Filamin-B levels. The work demonstrates that proteome integration at the single glomerulus and the individual organism levels can link \u201comics\u201d datasets to physiological function at high resolution.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Understanding podocyte-specific responses to injury at a systems level is difficult because injury leads to podocyte loss or an increase of extracellular matrix, altering glomerular cellular composition. Finding a window into early podocyte injury might help identify molecular pathways involved in the podocyte stress response.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>We developed an approach to apply proteome analysis to very small samples of purified podocyte fractions. To examine podocytes in early disease states in FSGS mouse models, we used podocyte fractions isolated from individual mice after chemical induction of glomerular disease (with Doxorubicin or LPS). We also applied single-glomerular proteome analysis to tissue from patients with FSGS.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      Transcriptome and proteome analysis of glomeruli from patients with FSGS revealed an underrepresentation of podocyte-specific genes and proteins in late-stage disease. Proteome analysis of purified podocyte fractions from FSGS mouse models showed an early stress response that includes perturbations of metabolic, mechanical, and proteostasis proteins. Additional analysis revealed a high correlation between the amount of proteinuria and expression levels of the mechanosensor protein Filamin-B. Increased expression of Filamin-B in podocytes in biopsy samples from patients with FSGS, in single glomeruli from proteinuric rats, and in podocytes undergoing mechanical stress suggests that this protein has a role in detrimental stress responses. In\n                      <jats:italic toggle=\"yes\">Drosophila<\/jats:italic>\n                      , nephrocytes with reduced filamin homolog Cher displayed altered filtration capacity, but exhibited no change in slit diaphragm structure.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>We identified conserved mechanisms of the podocyte stress response through ultrasensitive proteome analysis of human glomerular FSGS tissue and purified native mouse podocytes during early disease stages. This approach enables systematic comparisons of large-scale proteomics data and phenotype-to-protein correlation.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1681\/asn.2019030312","type":"journal-article","created":{"date-parts":[[2020,2,11]],"date-time":"2020-02-11T21:39:55Z","timestamp":1581457195000},"page":"544-559","source":"Crossref","is-referenced-by-count":35,"title":["Proteome Analysis of Isolated Podocytes Reveals Stress Responses in Glomerular Sclerosis"],"prefix":"10.1681","volume":"31","author":[{"given":"Sybille","family":"Koehler","sequence":"first","affiliation":[{"name":"Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany;"},{"name":"Biomedical Sciences, University of Edinburgh, Edinburgh, Scotland, United Kingdom;"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alexander","family":"Kuczkowski","sequence":"additional","affiliation":[{"name":"Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany;"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lucas","family":"Kuehne","sequence":"additional","affiliation":[{"name":"Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany;"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6586-7129","authenticated-orcid":false,"given":"Christian","family":"J\u00fcngst","sequence":"additional","affiliation":[{"name":"Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne, Germany;"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8698-5389","authenticated-orcid":false,"given":"Martin","family":"Hoehne","sequence":"additional","affiliation":[{"name":"Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany;"},{"name":"Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne, Germany;"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Florian","family":"Grahammer","sequence":"additional","affiliation":[{"name":"III. Department of Medicine, University Medical Center Hamburg-Eppendorf, Eppendorf, Hamburg, Germany;"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8578-3443","authenticated-orcid":false,"given":"Sean","family":"Eddy","sequence":"additional","affiliation":[{"name":"Division of Nephrology, Department of Internal Medicine, and"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4064-0582","authenticated-orcid":false,"given":"Matthias","family":"Kretzler","sequence":"additional","affiliation":[{"name":"Division of Nephrology, Department of Internal Medicine, and"},{"name":"Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, Michigan;"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bodo B.","family":"Beck","sequence":"additional","affiliation":[{"name":"Department of Human Genetics, University Hospital Cologne, Cologne, Germany;"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"J\u00f6rg","family":"H\u00f6hfeld","sequence":"additional","affiliation":[{"name":"Cell Biology, University of Bonn, Bonn, Germany; and"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bernhard","family":"Schermer","sequence":"additional","affiliation":[{"name":"Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany;"},{"name":"Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne, Germany;"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Thomas","family":"Benzing","sequence":"additional","affiliation":[{"name":"Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany;"},{"name":"Cologne 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