title: DENEDDYLASE1 Protein Counters Automodification of Neddylating Enzymes to Maintain NEDD8 Protein Homeostasis inArabidopsis.
abstract: In eukaryotes, the conjugation of the ubiquitin-like protein NEDD8 onto protein targets is an important post-translational modification. The best understood neddylation targets are the cullins, scaffold subunits of E3 ubiquitin ligases, where neddylation as well as deneddylation, facilitated by the protease activity of the CSN (COP9signalosome), are required to control ubiquitin ligase assembly, function, and ultimately substrate degradation. Little is known about the role of other deneddylating enzymes besides CSN and the role of neddylation and deneddylation of their substrates. We previously characterizedArabidopsis thalianamutants with defects in the conserved NEDD8-specific proteaseDEN1(DENEDDYLASE1). These mutants display only subtle growth phenotypes despite the strong accumulation of a broad range of neddylated proteins. Specifically, we identified AXR1 (AUXIN-RESISTANT1), a subunit of the heterodimeric NAE (E1 NEDD8-ACTIVATING ENZYME), as highly neddylated inden1mutants. Here, we examined the mechanism and consequences of AXR1 neddylation in more detail. We find that AXR1 as well as other neddylation enzymes are autoneddylated at multiple lysines. NAE autoneddylation can be linked to reduced NCE (E2 NEDD8-CONJUGATING ENZYME) NEDD8 thioester levels, either by critically reducing the pool of free NEDD8 or by reducing NAE activity.In planta, increasingNEDD8gene dosage is sufficient to suppressden1mutant phenotypes. We therefore suggest that DEN1 serves to recover diverted NEDD8 moieties from autoneddylated NAE subunits, and possibly also other neddylated proteins, to maintain NEDD8 pathway activity toward other NEDD8-dependent processes such as cullin E3 ligase regulation.
authors.affiliation: From the Chair of Plant Systems Biology, Technische Universität München, Emil-Ramann-Strasse 8 and.|the Chair of Proteomics and Bioanalytics, Technische Universität München, Emil-Erlenmeyer-Forum 5, 85354 Freising, Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Mergner
authors.affiliation: the Chair of Proteomics and Bioanalytics, Technische Universität München, Emil-Erlenmeyer-Forum 5, 85354 Freising, Germany.
authors.forename: Bernhard
authors.identifier: 0000-0002-9094-1677
authors.initials: B
authors.lastname: Kuster
authors.affiliation: From the Chair of Plant Systems Biology, Technische Universität München, Emil-Ramann-Strasse 8 and claus.schwechheimer@wzw.tum.de.
authors.forename: Claus
authors.identifier: 0000-0003-0269-2330
authors.initials: C
authors.lastname: Schwechheimer
mesh_terms: D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism*; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D004952:Esters / Q000737:chemistry; D018628:Gene Dosage; D006801:Humans; D008239:Lysine / Q000737:chemistry; D013058:Mass Spectrometry; D009154:Mutation; D010641:Phenotype; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000378:metabolism; D025801:Ubiquitin / Q000737:chemistry; D044764:Ubiquitin-Activating Enzymes / Q000378:metabolism; D014452:Ubiquitins / Q000235:genetics* / Q000378:metabolism*
chemical_list: C081977:AXR1 protein, Arabidopsis; D029681:Arabidopsis Proteins; C000620982:DEN1 protein, Arabidopsis; C112755:ECR1 protein, Arabidopsis; D004952:Esters; C513170:RUB1 protein, Arabidopsis; D011994:Recombinant Proteins; D025801:Ubiquitin; D014452:Ubiquitins; D010450:Endopeptidases; D044764:Ubiquitin-Activating Enzymes; C491802:NAE protein, human; D008239:Lysine
keywords: Arabidopsis; Arabidopsis thaliana; deubiquitylation (deubiquitination); ubiquitin; ubiquitin-dependent protease; ubiquitylation (ubiquitination)