An Eight Amino Acid Segment Controls Oligomerization and Preferred Conformation of the two Non-visual Arrestins

[Display omitted] •In the presence of IP6 arrestin-2 and arrestin-3 form different oligomers.•IP6 activates arrestin-3, but does not activate arrestin-2.•An eight-residue insertion in arrestin-2 controls differential oligomerization. G protein coupled receptors signal through G proteins or arrestins. A long-standing mystery in the field is why vertebrates have two non-visual arrestins, arrestin-2 and arrestin-3. These isoforms are ~75% identical and 85% similar; each binds numerous receptors, and appear to have many redundant functions, as demonstrated by studies of knockout mice. We previously showed that arrestin-3 can be activated by inositol-hexakisphosphate (IP6). IP6 interacts with the receptor-binding surface of arrestin-3, induces arrestin-3 oligomerization, and this oligomer stabilizes the active conformation of arrestin-3. Here, we compared the impact of IP6 on oligomerization and conformational equilibrium of the highly homologous arrestin-2 and arrestin-3 and found that these two isoforms are regulated differently. In the presence of IP6, arrestin-2 forms “infinite” chains, where each promoter remains in the basal conformation. In contrast, full length and truncated arrestin-3 form trimers and higher-order oligomers in the presence of IP6; we showed previously that trimeric state induces arrestin-3 activation (Chen et al., 2017). Thus, in response to IP6, the two non-visual arrestins oligomerize in different ways in distinct conformations. We identified an insertion of eight residues that is conserved across arrestin-2 homologs, but absent in arrestin-3 that likely accounts for the differences in the IP6 effect. Because IP6 is ubiquitously present in cells, this suggests physiological consequences, including differences in arrestin-2/3 trafficking and JNK3 activation. The functional differences between two non-visual arrestins are in part determined by distinct modes of their oligomerization. The mode of oligomerization might regulate the function of other signaling proteins.