RIC-3 expression and splicing regulate nAChR functional expression

Abstract

BackgroundThe nicotinic acetylcholine receptors form a large and diverse family of acetylcholine gated ion channels having diverse roles in the central nervous system. Maturation of nicotinic acetylcholine receptors is a complex and inefficient process requiring assistance from multiple cellular factors including RIC-3, a functionally conserved endoplasmic reticulum-resident protein and nicotinic acetylcholine receptor-specific chaperone. In mammals and in Drosophila melanogaster RIC-3 is alternatively spliced to produce multiple isoforms.ResultsWe used electrophysiological analysis in Xenopus laevis oocytes, in situ hybridization, and quantitative real-time polymerase chain reaction assays to investigate regulation of RIC-3’s expression and splicing and its effects on the expression of three major neuronal nicotinic acetylcholine receptors. We found that RIC-3 expression level and splicing affect nicotinic acetylcholine receptor functional expression and that two conserved RIC-3 isoforms express in the brain differentially. Moreover, in immune cells RIC-3 expression and splicing are regulated by inflammatory signals.ConclusionsRegulation of expression level and splicing of RIC-3 in brain and in immune cells following inflammation enables regulation of nicotinic acetylcholine receptor functional expression. Specifically, in immune cells such regulation via effects on α7 nicotinic acetylcholine receptor, known to function in the cholinergic anti-inflammatory pathway, may have a role in neuroinflammatory diseases.Electronic supplementary materialThe online version of this article (doi:10.1186/s13041-016-0231-5) contains supplementary material, which is available to authorized users.

RIC-3, a potential target for regulating cholinergic signaling and inflammation

“…This together with the known role of RIC-3 in α7 nAChR maturation suggests that RIC3 variants may affect MS via their effects on α7 nAChR maturation in immune cells. Importantly, we demonstrated that ric3 expression in immune cells is dynamically regulated following immune activation 31 . Moreover, recent unpublished results show that ric3 expression is transiently reduced in splenocytes and is gradually reduced in spinal cords during EAE progression (Yael BenDavid, unpublished).…”

“…. Recently we showed that these opposite effects on nAChR expression can be achieved using different RIC-3-toreceptor ratios, and that sensitivity of nAChR maturation to this ratio depends on the identity of the nAChR and on the specific RIC-3 isoform (different isoforms are produced by alternative splicing, see below) 31 . Second, RIC-3 was shown to affect different aspects of nAChR maturation depending on identity of the receptor's subunits.…”

“…First, RIC-3 was shown to have either positive or negative effects on co-expressed receptors depending on the RIC-3-to-receptor ratio 31 . Thus mechanisms regulating RIC-3 expression or stability are likely to regulate cholinergic signaling via nAChRs.…”

“…One conserved alternatively spliced RIC-3 isoform, lacking the conserved C-terminal coiled-coil domain, is defective for either the positive or negative effects of RIC-3 in a nAChR subtype dependent manner 31 .…”

Regulation of RIC-3 and of nAChR expression

“…This regulation of RIC-3 expression and splicing is likely to have functional implications as electrophysiology results, summarized above, show either positive or negative effects of RIC- 3 on nAChR expression, depending on RIC-3 expression level, specific RIC-3 isoform, and specific nAChR [5]. …”