Vsevolod V. Gurevich

Arrestins were discovered for their role in homologous desensitization of G‐protein‐coupled receptors (GPCRs). Later non‐visual arrestins were shown to regulate several signaling pathways. Some of these pathways require arrestin binding to GPCRs, the regulation of others is receptor independent. Here, we demonstrate that arrestin‐3 binds the E3 ubiquitin ligase parkin via multiple sites, preferentially interacting with its RING0 domain. Identification of the parkin domains involved suggests that arrestin‐3 likely relieves parkin autoinhibition and/or stabilizes the enzymatically active “open” conformation of parkin. Arrestin‐3 binding enhances ubiquitination by parkin of the mitochondrial protein mitofusin‐1 and facilitates parkin‐mediated mitophagy in HeLa cells. Furthermore, arrestin‐3 and its mutant with enhanced parkin binding rescue mitofusin‐1 ubiquitination and mitophagy in the presence of the Parkinson's disease‐associated R275W parkin mutant, which is defective in both functions. Thus, modulation of parkin activity via arrestin‐3 might be a novel strategy of anti‐parkinsonian therapy.

Arrestins bind active phosphorylated G protein-coupled receptors (GPCRs). Among the four mammalian subtypes, only arrestin-3 facilitates the activation of JNK3 in cells. In available structures, Lys-295 in the lariat loop of arrestin-3 and its homologue Lys-294 in arrestin-2 directly interact with the activator-attached phosphates. We compared the roles of arrestin-3 conformational equilibrium and Lys-295 in GPCR binding and JNK3 activation. Several mutants with enhanced ability to bind GPCRs showed much lower activity towards JNK3, whereas a mutant that does not bind GPCRs was more active. The subcellular distribution of mutants did not correlate with GPCR recruitment or JNK3 activation. Charge neutralization and reversal mutations of Lys-295 differentially affected receptor binding on different backgrounds but had virtually no effect on JNK3 activation. Thus, GPCR binding and arrestin-3-assisted JNK3 activation have distinct structural requirements, suggesting that facilitation of JNK3 activation is the function of arrestin-3 that is not bound to a GPCR.