title: SUMO E3 ligases are expressed in the retina and regulate SUMOylation of the metabotropic glutamate receptor 8b.
abstract: The central nervous system regulates neuronal excitability by macromolecular signalling complexes that consist of functionally related proteins, including neurotransmitter receptors, enzymes and scaffolds. The composition of these signal complexes is regulated by post-translational modifications, such as phosphorylation and SUMOylation (SUMO is small ubiquitin-related modifier). In the present study, we searched for proteins interacting with the intracellular C-termini of the metabotropic glutamate receptors mGluR8a and mGluR8b and identified proteins of the SUMOylation and NEDDylation machinery. The SUMO E3 ligases Pias1 [Pias is protein inhibitor of activated STAT (signal transducer and activator of transcription)] and Pias3L interacted strongly with mGluR8b, and were co-localized with the E2-conjugating Ubc9, SUMO1 and mGluR8b in cell bodies present in the ganglion cell layer of the mammalian retina. SUMO1 conjugation of Lys882, present in a bona fide consensus sequence for SUMOylation (VKSE) in the mGluR8b C-terminus, was enhanced by addition of Pias1, consistent with an interaction between both proteins. Mutation of Lys882 to arginine reduced, but did not abolish, mGluR8b SUMOylation. Co-mutating a second lysine residue (Lys903) located in the mGluR8b isoform-specific C-terminus largely prevented SUMO1 conjugation by Ubc9. Modelling studies suggested that Lys903 contacts Ubc9 and thus is part of the non-canonical SUMOylation site VKSG. In summary, the results of the present study show in vivo SUMOylation of the complete mGluR8b and co-localize proteins of the SUMOylation machinery in the retina.
authors.affiliation: Institut für Biochemie (Emil-Fischer-Zentrum), Friedrich-Alexander-Universität Erlangen-Nürnberg, 91045 Erlangen, Germany.
authors.forename: Eva
authors.initials: E
authors.lastname: Dütting
authors.forename: Nadja
authors.initials: N
authors.lastname: Schröder-Kress
authors.forename: Heinrich
authors.initials: H
authors.lastname: Sticht
authors.forename: Ralf
authors.initials: R
authors.lastname: Enz
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D020134:Catalytic Domain / Q000235:genetics; D002478:Cells, Cultured; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D051379:Mice; D008954:Models, Biological; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050828:Protein Inhibitors of Activated STAT / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D054730:Protein Interaction Domains and Motifs / Q000235:genetics; D020033:Protein Isoforms / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000235:genetics; D020836:Protein Structure, Quaternary; D051381:Rats; D018094:Receptors, Metabotropic Glutamate / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012160:Retina / Q000201:enzymology / Q000378:metabolism*; D025842:SUMO-1 Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D058207:Sumoylation / Q000235:genetics*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism
chemical_list: D050828:Protein Inhibitors of Activated STAT; D020033:Protein Isoforms; D018094:Receptors, Metabotropic Glutamate; D025842:SUMO-1 Protein; C111435:metabotropic glutamate receptor 8; D044767:Ubiquitin-Protein Ligases