The posttranslational addition of small ubiquitin-like modifier (SUMO) is an essential protein modification in plants that provides protection against numerous environmental challenges. Ligation is accomplished by a small set of SUMO ligases, with the SAP-MIZ domain-containing SIZ1 and METHYL METHANESULFONATE-SENSITIVE21 (MMS21) ligases having critical roles in stress protection and DNA endoreduplication/repair, respectively. To help identify their corresponding targets in , we used and mutants for proteomic analyses of SUMOylated proteins enriched via an engineered SUMO1 isoform suitable for mass spectrometric studies.
View Article and Find Full Text PDFMethods Mol Biol
January 2018
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.
View Article and Find Full Text PDFIn response to abiotic and biotic challenges, plants rapidly attach small ubiquitin-related modifier (SUMO) to a large collection of nuclear proteins, with studies in Arabidopsis (Arabidopsis thaliana) linking SUMOylation to stress tolerance via its modification of factors involved in chromatin and RNA dynamics. Despite this importance, little is known about SUMOylation in crop species. Here, we describe the plant SUMO system at the phylogenetic, biochemical, and transcriptional levels with a focus on maize (Zea mays).
View Article and Find Full Text PDFThe stress-induced attachment of small ubiquitin-like modifier (SUMO) to a diverse collection of nuclear proteins regulating chromatin architecture, transcription, and RNA biology has been implicated in protecting plants and animals against numerous environmental challenges. In order to better understand stress-induced SUMOylation, we combined stringent purification of SUMO conjugates with isobaric tag for relative and absolute quantification mass spectrometry and an advanced method to adjust for sample-to-sample variation so as to study quantitatively the SUMOylation dynamics of intact Arabidopsis seedlings subjected to stress. Inspection of 172 SUMO substrates during and after heat shock (37 °C) revealed that stress mostly increases the abundance of existing conjugates, as opposed to modifying new targets.
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