title: Purification of SUMO Conjugates from Arabidopsis for Mass Spectrometry Analysis.
abstract: The posttranslational modification of proteins with small ubiquitin-related modifier (SUMO) is a rapid, robust, and reversible mechanism that impacts a host of eukaryotic processes important to both normal cellular functions and survival during various abiotic and biotic challenges. Essential to defining the breadth of events impacted by SUMOylation is the development of full catalogues of protein targets. Here, we describe a stringent affinity method to purify native SUMO conjugates from the model plant Arabidopsis thaliana based on the expression of modified SUMOs bearing epitope tags. When combined with standard and quantitative mass spectrometric methods, deep datasets of SUMOylated proteins can be acquired. Functional analysis of these lists links SUMO to numerous regulatory events, with an emphasis on those associated with transcription, DNA replication and repair, and chromatin assembly/accessibility.
authors.affiliation: Department of Genetics, University of Wisconsin, 425-G Henry Mall, Madison, Wisconsin, 53706, USA.|Department of Biology, Washington University in St. Louis, Campus Box 1137, One Brookings Drive, St. Louis, Missouri, 63130, USA.
authors.forename: Thérèse C
authors.initials: TC
authors.lastname: Rytz
authors.affiliation: Department of Genetics, University of Wisconsin, 425-G Henry Mall, Madison, Wisconsin, 53706, USA.
authors.forename: Marcus J
authors.initials: MJ
authors.lastname: Miller
authors.affiliation: Department of Genetics, University of Wisconsin, 425-G Henry Mall, Madison, Wisconsin, 53706, USA. rdvierstra@wustl.edu.|Department of Biology, Washington University in St. Louis, Campus Box 1137, One Brookings Drive, St. Louis, Missouri, 63130, USA. rdvierstra@wustl.edu.
authors.forename: Richard D
authors.initials: RD
authors.lastname: Vierstra
mesh_terms: D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000737:chemistry; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D002843:Chromatin / Q000737:chemistry* / Q000378:metabolism; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism; D000939:Epitopes / Q000737:chemistry / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors / Q000737:chemistry / Q000378:metabolism; D018796:Immunoconjugates / Q000737:chemistry / Q000302:isolation & purification*; D047468:Immunoprecipitation; D008025:Ligases / Q000235:genetics / Q000378:metabolism; D013058:Mass Spectrometry; D020033:Protein Isoforms / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D058207:Sumoylation; D014158:Transcription, Genetic
chemical_list: D000911:Antibodies, Monoclonal; D029681:Arabidopsis Proteins; D002843:Chromatin; D000939:Epitopes; D018796:Immunoconjugates; D020033:Protein Isoforms; D011994:Recombinant Proteins; C477514:SCE1a protein, Arabidopsis; C477512:SUM1 protein, Arabidopsis; D003546:Cysteine Endopeptidases; C557297:ESD4 protein, Arabidopsis; D008025:Ligases; C502552:SIZ1 protein, Arabidopsis
keywords: Affinity purification; Arabidopsis thaliana; Mass spectrometry; Plants; SUMO; Small ubiquitin-like modifier