title: The Arabidopsis Iron-Sulfur Protein GRXS17 is a Target of the Ubiquitin E3 Ligases RGLG3 and RGLG4.
abstract: The stability of signaling proteins in eukaryotes is often controlled by post-translational modifiers. For polyubiquitination, specificity is assured by E3 ubiquitin ligases. Although plant genomes encode hundreds of E3 ligases, only few targets are known, even in the model Arabidopsis thaliana. Here, we identified the monothiol glutaredoxin GRXS17 as a substrate of the Arabidopsis E3 ubiquitin ligases RING DOMAIN LIGASE 3 (RGLG3) and RGLG4 using a substrate trapping approach involving tandem affinity purification of RING-dead versions. Simultaneously, we used a ubiquitin-conjugating enzym (UBC) panel screen to pinpoint UBC30 as a cognate E2 UBC capable of interacting with RGLG3 and RGLG4 and mediating auto-ubiquitination of RGLG3 and ubiquitination of GRXS17 in vitro. Accordingly, GRXS17 is ubiquitinated and degraded in an RGLG3- and RGLG4-dependent manner in planta. The truncated hemoglobin GLB3 also interacted with RGLG3 and RGLG4 but appeared to obstruct RGLG3 ubiquitination activity rather than being its substrate. Our results suggest that the RGLG family is intimately linked to the essential element iron.
authors: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052 Gent, Belgium Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052 Gent, Belgium These authors contributed equally to this work.
authors: Astrid
authors: A
authors: Nagels Durand
authors: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052 Gent, Belgium Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052 Gent, Belgium These authors contributed equally to this work.
authors: Sabrina
authors: S
authors: Iñigo
authors: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052 Gent, Belgium Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052 Gent, Belgium.
authors: Andrés
authors: A
authors: Ritter
authors: Plant Molecular Genetics Department, National Centre for Biotechnology (CNB), Consejo Superior de Investigaciones Científicas (CSIC), Campus Universidad Autónoma, Madrid, Spain.
authors: Elisa
authors: E
authors: Iniesto
authors: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052 Gent, Belgium Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052 Gent, Belgium.
authors: Rebecca
authors: R
authors: De Clercq
authors: Medical Biotechnology Center, VIB, B-9000 Ghent, Belgium Department of Biochemistry, Ghent University, B-9000 Ghent, Belgium.
authors: An
authors: A
authors: Staes
authors: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052 Gent, Belgium Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052 Gent, Belgium.
authors: Jelle
authors: J
authors: Van Leene
authors: Plant Molecular Genetics Department, National Centre for Biotechnology (CNB), Consejo Superior de Investigaciones Científicas (CSIC), Campus Universidad Autónoma, Madrid, Spain.
authors: Vicente
authors: V
authors: Rubio
authors: Medical Biotechnology Center, VIB, B-9000 Ghent, Belgium Department of Biochemistry, Ghent University, B-9000 Ghent, Belgium.
authors: Kris
authors: K
authors: Gevaert
authors: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052 Gent, Belgium Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052 Gent, Belgium.
authors: Geert
authors: G
authors: De Jaeger
authors: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052 Gent, Belgium Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052 Gent, Belgium These authors contributed equally to this work.
authors: Laurens
authors: L
authors: Pauwels
authors: Department of Plant Systems Biology, VIB, Technologiepark 927, B-9052 Gent, Belgium Department of Plant Biotechnology and Bioinformatics, Ghent University, Technologiepark 927, B-9052 Gent, Belgium These authors contributed equally to this work. algoo@psb.vib-ugent.be.
authors: Alain
authors: A
authors: Goossens
mesh_terms: D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D003517:Cyclopentanes / Q000378:metabolism; D054477:Glutaredoxins / Q000235:genetics / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000378:metabolism; D008025:Ligases / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D054883:Oxylipins / Q000378:metabolism; D030821:Plants, Genetically Modified; D025821:Polyubiquitin / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques; D044763:Ubiquitin-Conjugating Enzymes / Q000235:genetics / Q000378:metabolism; D054875:Ubiquitination
chemical_list: D029681:Arabidopsis Proteins; D003517:Cyclopentanes; C000604704:GRXS17 protein, Arabidopsis; D054477:Glutaredoxins; D007506:Iron-Sulfur Proteins; D054883:Oxylipins; D025821:Polyubiquitin; C011006:jasmonic acid; C576451:UBC32 protein, Arabidopsis; D044763:Ubiquitin-Conjugating Enzymes; D046988:Proteasome Endopeptidase Complex; D008025:Ligases; C000604772:RGLG3 protein, Arabidopsis; C000604773:RGLG4 protein, Arabidopsis
keywords: Arabidopsis; E3 ubiquitin ligase; Glutaredoxin; Post-translational modification; Thioredoxin; Ubiquitination