High-Confidence Interactome for RNF41 Built on Multiple Orthogonal Assays

Ring
finger protein 41 (RNF41) is an E3 ubiquitin ligase involved
in the ubiquitination and degradation of many proteins including ErbB3
receptors, BIRC6, and parkin. Next to this, RNF41 regulates the intracellular
trafficking of certain JAK2-associated cytokine receptors by ubiquitinating
and suppressing USP8, which, in turn, destabilizes the ESCRT-0 complex.
To further elucidate the function of RNF41 we used different orthogonal
approaches to reveal the RNF41 protein complex: affinity purification–mass
spectrometry, BioID, and Virotrap. We combined these results with
known data sets for RNF41 obtained with microarray MAPPIT and Y2H
screens. This way, we establish a comprehensive high-resolution interactome
network comprising 175 candidate protein partners. To remove potential
methodological artifacts from this network, we distilled the data
into a high-confidence interactome map by retaining a total of 19
protein hits identified in two or more of the orthogonal methods.
AP2S1, a novel RNF41 interaction partner, was selected from this high-confidence
interactome for further functional validation. We reveal a role for
AP2S1 in leptin and LIF receptor signaling and show that RNF41 stabilizes
and relocates AP2S1.