Top Highlights

• RNF213 features six AAA-ATPase modules, two of which are active, a RING E3 ligase domain, a recently discovered “RZ” E3 domain, and large unstructured regions23-27. In an AAA- ATPase- and ISG15-dependent manner, RNF213 can oligomerize into a hexameric, ~3.6mDa complex23,25. RNF213 localizes on lipid droplets28 (also dependent on AAA-ATPase activity...
• We generated an RNF213-TurboID fusion protein with the goal of identifying RNF213 binding partners by proximity ligation31 (see below), labelled interacting proteins with biotin, enriched these proteins on streptavidin beads, performed on-bead tryptic digestion, and analyzed the peptide mixture by mass spectrometry (MS). This analysis revealed evid...
• We therefore tested the hypothesis that phosphorylation affects RNF213 oligomerization by co-transfecting expression vectors for 3x- FLAG-RNF213 or 3x-FLAG-RNF213Y1275F and GFP-RNF213 and performing anti-FLAG immunoprecipitations.
• B-catalyzed dephosphorylation. PTP1B deficiency/inhibition markedly increases overall cellular ubiquitylation, dependent on RNF21322. Y1275 is distant from the RING or RZ domains in the monomeric RNF213 structure23, arguing against direct regul

Domain