Human genetic variants disrupt RGS14 nuclear shuttling and regulation of LTP in hippocampal neurons

Abstract

The human genome contains vast genetic diversity as naturally occurring coding variants, yet the impact of these variants on protein function and physiology is poorly understood. RGS14 is a multifunctional signaling protein that suppresses synaptic plasticity in dendritic spines of hippocampal neurons. RGS14 also is a nucleocytoplasmic shuttling protein, suggesting that balanced nuclear import/export and dendritic spine localization are essential for RGS14 functions. We identified genetic variants L505R (LR) and R507Q (RQ) located within the nuclear export sequence (NES) of human RGS14. Here we report that RGS14 encoding LR or RQ profoundly impacts protein functions in hippocampal neurons. RGS14 membrane localization is regulated by binding Gαi-GDP, whereas RGS14 nuclear export is regulated by Exportin 1 (XPO1). Remarkably, LR and RQ variants disrupt RGS14 binding to Gαi1-GDP and XPO1, nucleocytoplasmic equilibrium, and capacity to inhibit LTP. Variant LR accumulates irreversibly in the nucleus, preventing RGS14 binding to Gαi1, localization to dendritic spines, and inhibitory actions on LTP induction, while variant RQ exhibits a mixed phenotype. When introduced into mice by CRISPR/Cas9, RGS14-LR protein expression was detected predominantly in the nuclei of neurons within hippocampus, central amygdala, piriform cortex, and striatum, brain regions associated with learning and synaptic plasticity. Whereas mice completely lacking RGS14 exhibit enhanced spatial learning, mice carrying variant LR exhibit normal spatial learning, suggesting that RGS14 may have distinct functions in the nucleus independent from those in dendrites and spines. These findings show that naturally occurring genetic variants can profoundly alter normal protein function, impacting physiology in unexpected ways.

RGS14 Regulation of Post-Synaptic Signaling and Spine Plasticity in Brain

“…However, RGS14 suppression of LTP in CA2 depends, in part, on PKA inhibition [79], suggesting other factors are also important. Consistent with this idea, our most recent report demonstrates that the RGS domain of RGS14 is not essential for its inhibitory effects on LTP [19]. Therefore, RGS14 inhibits synaptic plasticity and likely PKA in a manner independent of its effect on Gi/o-cAMP signaling.…”

“…In the adult brain, RGS14 is expressed in the hippocampus, piriform cortex, entorhinal cortex, layers II, III, and V of the neocortex, the anterior olfactory nucleus, and the orbital cortex [27]. More recently, we reported RGS14 expression in neurons of the central amygdala and the striatum [19]. In primates, including rhesus macaques and post-mortem tissue from humans [26], we find RGS14 protein expression in the caudate nucleus, putamen, and globus pallidus.…”

“…An additional population of presumably excitatory RGS14-containing inputs of yet-unknown origin is also found in the substantia nigra pars reticulata. Another region enriched with RGS14 is the amygdala, in which expression is found in putative pro-jection neurons of the basomedial, basolateral, and centrolateral nuclei [19,56]. Projections from the central nucleus of the amygdala may comprise the unidentified RGS14-positive innervation in the substantia nigra, though further study is required.…”

“…Despite not being essential for RGS14's effects on plasticity [19], RGS control of Gi/o signaling is still important in the hippocampus and elsewhere. A number of Gi/o-coupled GPCRs are highly expressed in area CA2 [66].…”

“…With respect to RGS14 effects on LTP and synaptic plasticity, the GPR motif is essential for RGS14's inhibitory actions in hippocampal neurons [19]. A mutation in the GPR motif of RGS14, which prevents binding to Gαi1/3, abolishes RGS14's capacity to inhibit LTP in hippocampal slices.…”