Speculation on How RIC-3 and Other Chaperones Facilitate α7 Nicotinic Receptor Folding and Assembly

Abstract

The process of how multimeric transmembrane proteins fold and assemble in the endoplasmic reticulum is not well understood. The alpha7 nicotinic receptor (α7 nAChR) is a good model for multimeric protein assembly since it has at least two independent and specialized chaperones: Resistance to Inhibitors of Cholinesterase 3 (RIC-3) and Nicotinic Acetylcholine Receptor Regulator (NACHO). Recent cryo-EM and NMR data revealed structural features of α7 nAChRs. A ser-ala-pro (SAP) motif precedes a structurally important but unique “latch” helix in α7 nAChRs. A sampling of α7 sequences suggests the SAP motif is conserved from C. elegans to humans, but the latch sequence is only conserved in vertebrates. How RIC-3 and NACHO facilitate receptor subunits folding into their final pentameric configuration is not known. The artificial intelligence program AlphaFold2 recently predicted structures for NACHO and RIC-3. NACHO is highly conserved in sequence and structure across species, but RIC-3 is not. This review ponders how different intrinsically disordered RIC-3 isoforms from C. elegans to humans interact with α7 nAChR subunits despite having little sequence homology across RIC-3 species. Two models from the literature about how RIC-3 assists α7 nAChR assembly are evaluated considering recent structural information about the receptor and its chaperones.

Unraveling the molecular interactions between α7 nicotinic receptor and a RIC3 variant associated with backward speech

“…If the misfolded proteins are α7, then this implicates a direct role for RIC3 in α7 folding as suggested by [ 17 ]. Given the disordered nature of RIC3, the localization of the exact interaction domain has proven to be challenging [ 41 ].…”

“…The exact mechanism by which RIC3 promotes α7 nAChR assembly is unknown [ 41 ] although a direct interaction is expected as α7 co-precipitates with RIC3 [ 20 , 42 ]. Wang et al suggested that each RIC3 protein associates with a single folded α7 subunit [ 17 ].…”

“…Each of these chaperones is thought to bind a distinct region of α7 [ 44 ] and it has been proposed that RIC3 binds between the M3 and M4 transmembrane domains [ 44 ]. This loop is largely disorganized but includes an MX helix and an MA helix, that runs into the M4 transmembrane domain [ 41 ]. Although studies have shown that this region is necessary for the effects of RIC3 [ 44 ] and that substitution of residues in the MA helix ablates RIC3 enhancement of assembly [ 25 ], structural modeling suggests that interactions in this region would block pentameric assembly [ 41 ].…”

Decoding alpha7 nicotinic receptor chaperone actions using chimeric proteins

“…Noviello et al (2021) using cryo-EM identified the conformational structures of the human α7 nAChRs and a C-terminal substructure involved in gating they called the "latch" [10]. Together with TM4, the "latch" may play an important role in RIC3 actions [10,19]. There is a highly conserved ser-ala-pro (SAP) motif located between TM4 and the "latch", suggesting that the SAP motif may have important effects on the folding and function of α7 nAChRs [19].…”

“…Together with TM4, the "latch" may play an important role in RIC3 actions [10,19]. There is a highly conserved ser-ala-pro (SAP) motif located between TM4 and the "latch", suggesting that the SAP motif may have important effects on the folding and function of α7 nAChRs [19]. Noviello et al also proposed the ICD has a higher proportion of disordered regions than the ECD and TMDs [10].…”