title: Identification of Targets and Interaction Partners of Arginyl-tRNA Protein Transferase in the Moss Physcomitrella patens.
abstract: Protein arginylation is a posttranslational modification of both N-terminal amino acids of proteins and sidechain carboxylates and can be crucial for viability and physiology in higher eukaryotes. The lack of arginylation causes severe developmental defects in moss, affects the low oxygen response in Arabidopsis thaliana and is embryo lethal in Drosophila and in mice. Although several studies investigated impact and function of the responsible enzyme, the arginyl-tRNA protein transferase (ATE) in plants, identification of arginylated proteins by mass spectrometry was not hitherto achieved. In the present study, we report the identification of targets and interaction partners of ATE in the model plant Physcomitrella patens by mass spectrometry, employing two different immuno-affinity strategies and a recently established transgenic ATE:GUS reporter line (Schuessele et al., 2016 New Phytol. , DOI: 10.1111/nph.13656). Here we use a commercially available antibody against the fused reporter protein (β-glucuronidase) to pull down ATE and its interacting proteins and validate its in vivo interaction with a class I small heatshock protein via Förster resonance energy transfer (FRET). Additionally, we apply and modify a method that already successfully identified arginylated proteins from mouse proteomes by using custom-made antibodies specific for N-terminal arginine. As a result, we identify four arginylated proteins from Physcomitrella patens with high confidence.Data are available via ProteomeXchange with identifier PXD003228 and PXD003232.
authors.affiliation: From the ‡Plant Biotechnology, Faculty of Biology, University of Freiburg, Schaenzlestr. 1, 79104 Freiburg, Germany;
authors.forename: Sebastian N W
authors.initials: SN
authors.lastname: Hoernstein
authors.affiliation: From the ‡Plant Biotechnology, Faculty of Biology, University of Freiburg, Schaenzlestr. 1, 79104 Freiburg, Germany;
authors.forename: Stefanie J
authors.initials: SJ
authors.lastname: Mueller
authors.affiliation: From the ‡Plant Biotechnology, Faculty of Biology, University of Freiburg, Schaenzlestr. 1, 79104 Freiburg, Germany;
authors.forename: Kathrin
authors.initials: K
authors.lastname: Fiedler
authors.affiliation: From the ‡Plant Biotechnology, Faculty of Biology, University of Freiburg, Schaenzlestr. 1, 79104 Freiburg, Germany;
authors.forename: Marc
authors.initials: M
authors.lastname: Schuelke
authors.affiliation: §Rudolf Virchow Center for Experimental Biomedicine, University of Wuerzburg, Josef-Schneider-Str. 2, 97080 Wuerzburg, Germany;
authors.forename: Jens T
authors.initials: JT
authors.lastname: Vanselow
authors.affiliation: From the ‡Plant Biotechnology, Faculty of Biology, University of Freiburg, Schaenzlestr. 1, 79104 Freiburg, Germany;
authors.forename: Christian
authors.initials: C
authors.lastname: Schuessele
authors.affiliation: From the ‡Plant Biotechnology, Faculty of Biology, University of Freiburg, Schaenzlestr. 1, 79104 Freiburg, Germany;
authors.forename: Daniel
authors.initials: D
authors.lastname: Lang
authors.affiliation: ¶ZBSA - Centre for Biological Systems Analysis, Life Imaging Center, University Freiburg, Habsburgerstr. 49, 79104 Freiburg, Germany; ‡‡BIOSS - Centre for Biological Signalling Studies, 79104 Freiburg, Germany.
authors.forename: Roland
authors.initials: R
authors.lastname: Nitschke
authors.affiliation: ‖Institute of Biology 3, Faculty of Biology, University of Freiburg, Schaenzlestr. 1, 79104 Freiburg, Germany;
authors.forename: Gabor L
authors.initials: GL
authors.lastname: Igloi
authors.affiliation: §Rudolf Virchow Center for Experimental Biomedicine, University of Wuerzburg, Josef-Schneider-Str. 2, 97080 Wuerzburg, Germany;
authors.forename: Andreas
authors.initials: A
authors.lastname: Schlosser
authors.affiliation: From the ‡Plant Biotechnology, Faculty of Biology, University of Freiburg, Schaenzlestr. 1, 79104 Freiburg, Germany; ¶ZBSA - Centre for Biological Systems Analysis, Life Imaging Center, University Freiburg, Habsburgerstr. 49, 79104 Freiburg, Germany; **FRIAS - Freiburg Institute for Advanced Studies, 79104 Freiburg, Germany; ‡‡BIOSS - Centre for Biological Signalling Studies, 79104 Freiburg, Germany ralf.reski@biologie.uni-freiburg.de.
authors.forename: Ralf
authors.initials: R
authors.lastname: Reski
mesh_terms: D019881:Aminoacyltransferases / Q000378:metabolism*; D000906:Antibodies / Q000378:metabolism; D019068:Bryopsida / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D013058:Mass Spectrometry; D010940:Plant Proteins / Q000737:chemistry / Q000378:metabolism*; D060066:Protein Interaction Maps; D040901:Proteomics / Q000379:methods
chemical_list: D000906:Antibodies; D010940:Plant Proteins; D019881:Aminoacyltransferases; C019727:arginyltransferase