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1. Proteomic and transcriptomic profiling of aerial organ development in Arabidopsis

Author

Mergner, Julia, Frejno, Martin, Messerer, Maxim, Lang, Daniel, Samaras, Patroklos, Wilhelm, Mathias, Mayer, Klaus F. X., Schwechheimer, Claus, and Kuster, Bernhard

Subjects
Statistics and Probability, Proteomics, Proteome, Gene Expression Profiling, fungi, Arabidopsis, food and beverages, Library and Information Sciences, Computer Science Applications, Education, lcsh:Q, Statistics, Probability and Uncertainty, Transcriptome, lcsh:Science, Information Systems
Abstract

Plant growth and development are regulated by a tightly controlled interplay between cell division, cell expansion and cell differentiation during the entire plant life cycle from seed germination to maturity and seed propagation. To explore some of the underlying molecular mechanisms in more detail, we selected different aerial tissue types of the model plant Arabidopsis thaliana , namely rosette leaf, flower and silique/seed and performed proteomic, phosphoproteomic and transcriptomic analyses of sequential growth stages using tandem mass tag-based mass spectrometry and RNA sequencing. With this exploratory multi-omics dataset, development dynamics of photosynthetic tissues can be investigated from different angles. As expected, we found progressive global expression changes between growth stages for all three omics types and often but not always corresponding expression patterns for individual genes on transcript, protein and phosphorylation site level. The biggest difference between proteomic- and transcriptomic-based expression information could be observed for seed samples. Proteomic and transcriptomic data is available via ProteomeXchange and ArrayExpress with the respective identifiers PXD018814 and E-MTAB-7978.

Published
2020

3. Challenges in Clinical Metaproteomics Highlighted by the Analysis of Acute Leukemia Patients with Gut Colonization by Multidrug-Resistant Enterobacteriaceae

Author

Rechenberger, Julia, Samaras, Patroklos, Jarzab, Anna, Behr, Juergen, Frejno, Martin, Djukovic, Ana, Sanz, Jaime, González-Barberá, Eva M., Salavert, Miguel, López-Hontangas, Jose Luis, Xavier, Karina B., Debrauwer, Laurent, Rolain, Jean-Marc, Sanz, Miguel, Garcia-Garcera, Marc, Wilhelm, Mathias, Ubeda, Carles, and Kuster, Bernhard

Subjects
ddc
Published
2019

4. Challenges in clinical metaproteomics highlighted by the analysis of acute leukemia patients with gut colonization by multidrug-resistant enterobacteriaceae

Author

Rechenberger, Julia, Samaras, Patroklos, Jarzab, Anna, Behr, Juergen, Frejno, Martin, Djukovic, Ana, Sanz, Jaime, Gonzalez-Barbera, Eva M., Salavert, Miguel, Luis Lopez-Hontangas, Jose, Xavier, Karina B., Debrauwer, Laurent, Rolain, Jean-Marc, Sanz, Miguel, Garcia-Garcera, Marc, Ubeda, Carles, Kuster, Bernhard, and Wilhelm, Mathias

Subjects
human gut microbiome, metaproteome, data analysis, mass spectrometry, proteomics, clinical proteomics, multi-omics, multidrug-resistant Enterobacteriaceae, Bioengineering, Ingénierie biomédicale
Abstract

The microbiome has a strong impact on human health and disease and is, therefore, increasingly studied in a clinical context. Metaproteomics is also attracting considerable attention, and such data can be efficiently generated today owing to improvements in mass spectrometry-based proteomics. As we will discuss in this study, there are still major challenges notably in data analysis that need to be overcome. Here, we analyzed 212 fecal samples from 56 hospitalized acute leukemia patients with multidrug-resistant Enterobactericeae (MRE) gut colonization using metagenomics and metaproteomics. This is one of the largest clinical metaproteomic studies to date, and the first metaproteomic study addressing the gut microbiome in MRE colonized acute leukemia patients. Based on this substantial data set, we discuss major current limitations in clinical metaproteomic data analysis to provide guidance to researchers in the field. Notably, the results show that public metagenome databases are incomplete and that sample-specific metagenomes improve results. Furthermore, biological variation is tremendous which challenges clinical study designs and argues that longitudinal measurements of individual patients are a valuable future addition to the analysis of patient cohorts.

Published
2019

5. Challenges in Clinical Metaproteomics Highlighted by the Analysis of Acute Leukemia Patients with Gut Colonization by Multidrug-Resistant Enterobacteriaceae

Author

Rechenberger, Julia, Samaras, Patroklos, Jarzab, Anna, Behr, Juergen, Frejno, Martin, Djukovic, Ana, Sanz, Jaime, González-Barberá, Eva, Salavert, Miguel, López-Hontangas, Jose Luis, Xavier, Karina, Debrauwer, Laurent, Rolain, Jean-Marc, Sanz, Miguel, Garcia-Garcerà, Marc, Wilhelm, Mathias, Ubeda, Carles, Kuster, Bernhard, Technische Universität Munchen - Université Technique de Munich [Munich, Allemagne] (TUM), Fundación para el Fomento de la Investigación Sanitaria y Biomédica de la Comunitat Valenciana [Espagne] (FISABIO), Hospital Universitari i Politècnic La Fe, Centro de Investigación Biomédica en Red de Cáncer (CIBERONC), Instituto Gulbenkian de Ciência [Oeiras] (IGC), Fundação Calouste Gulbenkian, Analyse de Xénobiotiques, Identification, Métabolisme (E20 Metatoul-AXIOM), ToxAlim (ToxAlim), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Ecole Nationale Vétérinaire de Toulouse (ENVT), Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Ecole d'Ingénieurs de Purpan (INPT - EI Purpan), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Recherche Agronomique (INRA)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Recherche Agronomique (INRA)-MetaToul-MetaboHUB, Génopole Toulouse Midi-Pyrénées [Auzeville] (GENOTOUL), Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Ecole Nationale Vétérinaire de Toulouse (ENVT), Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Génopole Toulouse Midi-Pyrénées [Auzeville] (GENOTOUL), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Institut Hospitalier Universitaire Méditerranée Infection (IHU Marseille), Université de Lausanne (UNIL), CIBER in Epidemiology and Public Health, BMBF FloraStopMRE grant [031L0089], Microbes évolution phylogénie et infections (MEPHI), Institut de Recherche pour le Développement (IRD)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Ecole d'Ingénieurs de Purpan (INPT - EI Purpan), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-MetaToul-MetaboHUB, Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Hospital Universitari i Politècnic La Fe = University and Polytechnic Hospital La Fe, MetaToul AXIOM (E20), Université de Toulouse (UT)-Université de Toulouse (UT)-Ecole Nationale Vétérinaire de Toulouse (ENVT), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université de Toulouse (UT)-Ecole d'Ingénieurs de Purpan (INP - PURPAN), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National de la Recherche Agronomique (INRA)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-MetaboHUB-MetaToul, MetaboHUB-Génopole Toulouse Midi-Pyrénées [Auzeville] (GENOTOUL), Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-MetaboHUB-Génopole Toulouse Midi-Pyrénées [Auzeville] (GENOTOUL), Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), and Université de Lausanne = University of Lausanne (UNIL)

Subjects
[SDV.MHEP.ME]Life Sciences [q-bio]/Human health and pathology/Emerging diseases, human gut microbiome, data analysis, lcsh:QR1-502, multi-omics, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, lcsh:Microbiology, Article, proteomics, [SDV.MHEP.CSC]Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, clinical proteomics, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, mass spectrometry, metaproteome, multidrug-resistant Enterobacteriaceae, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, [SDV.MP.PAR]Life Sciences [q-bio]/Microbiology and Parasitology/Parasitology
Abstract

International audience; The microbiome has a strong impact on human health and disease and is, therefore, increasingly studied in a clinical context. Metaproteomics is also attracting considerable attention, and such data can be efficiently generated today owing to improvements in mass spectrometry-based proteomics. As we will discuss in this study, there are still major challenges notably in data analysis that need to be overcome. Here, we analyzed 212 fecal samples from 56 hospitalized acute leukemia patients with multidrug-resistant Enterobactericeae (MRE) gut colonization using metagenomics and metaproteomics. This is one of the largest clinical metaproteomic studies to date, and the first metaproteomic study addressing the gut microbiome in MRE colonized acute leukemia patients. Based on this substantial data set, we discuss major current limitations in clinical metaproteomic data analysis to provide guidance to researchers in the field. Notably, the results show that public metagenome databases are incomplete and that sample-specific metagenomes improve results. Furthermore, biological variation is tremendous which challenges clinical study designs and argues that longitudinal measurements of individual patients are a valuable future addition to the analysis of patient cohorts.

Published
2019

6. Optimized Enrichment of Phosphoproteomes by Fe-IMAC Column Chromatography

Author

Ruprecht, Benjamin, Koch, Heiner, Domasinska, Petra, Frejno, Martin, Kuster, Bernhard, Lemeer, Simone, Walker, John M., Comai, Lucio, Katz, Jonathan E., Mallick, Parag, Sub Biomol.Mass Spectrometry & Proteom., and Biomolecular Mass Spectrometry and Proteomics

Subjects
Proteomics, Phosphorylation, Phosphocapture, LC-MS
Abstract

Phosphorylation is among the most important post-translational modifications of proteins and has numerous regulatory functions across all domains of life. However, phosphorylation is often substoichiometric, requiring selective and sensitive methods to enrich phosphorylated peptides from complex cellular digests. Various methods have been devised for this purpose and we have recently described a Fe-IMAC HPLC column chromatography setup which is capable of comprehensive, reproducible, and selective enrichment of phosphopeptides out of complex peptide mixtures. In contrast to other formats such as StageTips or batch incubations using TiO2 or Ti-IMAC beads, Fe-IMAC HPLC columns do not suffer from issues regarding incomplete phosphopeptide binding or elution and enrichment efficiency scales linearly with the amount of starting material. Here, we provide a step-by-step protocol for the entire phosphopeptide enrichment procedure including sample preparation (lysis, digestion, desalting), Fe-IMAC column chromatography (column setup, operation, charging), measurement by LC-MS/MS (nHPLC gradient, MS parameters) and data analysis (MaxQuant). To increase throughput, we have optimized several key steps such as the gradient time of the Fe-IMAC separation (15 min per enrichment), the number of consecutive enrichments possible between two chargings (>20) and the column recharging itself (90 %) identification of more than 10,000 unique phosphopeptides from 1 mg of HeLa digest within 2 h of measurement time (Q Exactive Plus).

Published
2017

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