Nuclear Targeting of an Endosomal E3 Ubiquitin Ligase

Abstract

Ring finger protein 13 (RNF13) is an E3 ubiquitin ligase embedded in endosome membranes. The protein undergoes constitutive post-translational proteolysis, making its detection difficult unless cells are incubated with a proteasome inhibitor to allow biosynthetic forms to accumulate. When cells were treated with phorbol 12-myristate 13-acetate (PMA), RNF13 avoided proteolysis. A similar stabilization was seen on ionomycin treatment of cells. Drug treatment stabilized both the full-length protein and a membrane-embedded C-terminal fragment generated following ectodomain shedding. Immunofluorescence staining revealed that PMA treatment caused the protein to accumulate in recycling endosomes, where it colocalized with transferrin receptor, and on the inner nuclear membrane, where it colocalized with lamin B. Expression of dominant-negative Rab11 inhibited nuclear localization, suggesting RNF13 was targeted to the inner nuclear membrane through recycling endosomes. New protein synthesis was necessary for this targeting. Nuclear localization was confirmed by immunoelectron microscopy and by purification of the inner nuclear membrane. Stress-induced transport of an endosomal protein to the inner nuclear membrane is a novel mechanism for introduction of regulatory proteins to the DNA environment. RNF13, with its ubiquitin ligase-active RING domain, has the potential to turn over key nuclear proteins in response to signals received at the plasma membrane.

The RING finger E3 ligase STRF1 is involved in membrane trafficking and modulates salt‐stress response in Arabidopsis thaliana

“…Indeed, other RING E3 ligases similar to STRF1 location have been identified in mammals. They include, among others, Tal1, which mediates Tsg101 ubiquitination and cargo sorting into vesicles (Amit et al, 2004) and RNF13, which localizes to the endosome membrane, and may take part in the ubiquitin-mediated endosome/lysosome recycling system (Bocock et al, 2009(Bocock et al, , 2010(Bocock et al, , 2011. Until now, only one E3 ligase from Arabidopsis, KEG (KEEP ON GOING), has been reported to localize to the TGN/EE, and to regulate endocytic trafficking and/or the formation of signaling complexes on TGN/EE vesicles during stress responses (Gu and Innes, 2011).…”

Comprehensively Surveying Structure and Function of RING Domains from Drosophila melanogaster

“…Traf6 also interacts with various protein kinases including IRAK1/IRAK, SRC and PKCzeta, which provides a link between distinct signaling pathways. In addition, RNF13 (NP_009213), an integral membrane-associated RING-E3, is targeted to the inner nuclear membrane through recycling endosomes, and has the potential to turn over key nuclear proteins in response to signals received at the plasma membrane [58].…”

The Role of the Transmembrane RING Finger Proteins in Cellular and Organelle Function

“…Recent studies have reported that RNF13 is present in the endosomal and lysosomal compartments, and the C-terminal region containing the RNF domain is released into the cytosol by proteolytic cleavage [125]. Activation of protein kinase C inhibits this cleavage and stabilizes the full-length RNF13 [127]. The stabilized RNF13 is then transported to the inner nuclear membrane via the recycling endosomes, thereby exposing its RNF domain to the nucleoplasm.…”

“…The stabilized RNF13 is then transported to the inner nuclear membrane via the recycling endosomes, thereby exposing its RNF domain to the nucleoplasm. RNF13 may modulate gene expression and signal transduction by mediating ubiquitination in response to various stimuli [35,127]. It is imperative to identify the RNF13 substrate proteins to obtain a better understanding of the physiological role and mechanism of action of RNF13.…”