Loss, Gain and Altered Function of GlyR α2 Subunit Mutations in Neurodevelopmental Disorders

Abstract

Glycine receptors (GlyRs) containing the α2 subunit govern cell fate, neuronal migration and synaptogenesis in the developing cortex and spinal cord. Rare missense variants and microdeletions in the X-linked GlyR α2 subunit gene (GLRA2) have been associated with human autism spectrum disorder (ASD), where they typically cause a loss-of-function via protein truncation, reduced cell-surface trafficking and/or reduced glycine sensitivity (e.g., GLRA2Δex8-9 and extracellular domain variants p.N109S and p.R126Q). However, the GlyR α2 missense variant p.R323L in the intracellular M3-M4 domain results in a gain-of-function characterized by slower synaptic decay times, longer duration active periods and increases in channel conductance. This study reports the functional characterization of four missense variants in GLRA2 associated with ASD or developmental disorders (p.V-22L, p.N38K, p.K213E, p.T269M) using a combination of bioinformatics, molecular dynamics simulations, cellular models of GlyR trafficking and electrophysiology in artificial synapses. The GlyR α2V–22L variant resulted in altered predicted signal peptide cleavage and a reduction in cell-surface expression, suggestive of a partial loss-of-function. Similarly, GlyR α2N38K homomers showed reduced cell-surface expression, a reduced affinity for glycine and a reduced magnitude of IPSCs in artificial synapses. By contrast, GlyR α2K213E homomers showed a slight reduction in cell-surface expression, but IPSCs were larger, with faster rise/decay times, suggesting a gain-of-function. Lastly, GlyR α2T269M homomers exhibited a high glycine sensitivity accompanied by a substantial leak current, suggestive of an altered function that could dramatically enhance glycinergic signaling. These results may explain the heterogeneity of clinical phenotypes associated with GLRA2 mutations and reveal that missense variants can result in a loss, gain or alteration of GlyR α2 function. In turn, these GlyR α2 missense variants are likely to either negatively or positively deregulate cortical progenitor homeostasis and neuronal migration in the developing brain, leading to changes in cognition, learning, and memory.

Age-Related microRNA Overexpression in Lafora Disease Male Mice Provides Links between Neuroinflammation and Oxidative Stress

“…Regarding Btg2, only a slight but significant decrease in the expression levels was registered in the Epm2a−/− mice. All the genes analyzed are somehow related to molecular pathways relevant to the pathophysiological context of LD: Nsun3 (NOP2/Sun RNA methyltransferase 3) encodes a mitochondrial methyltransferase related to encephalomyopathy and seizures [ 18 ]; Glra2 (glycine receptor alpha 2) encodes for a glycine receptor that has been predicted to influence cognition, learning, and memory in the developing brain [ 19 ]; Btg2 (BTG anti-proliferation factor 2) is an anti-proliferative gene that, interestingly, has been postulated as a neuroprotective gene in mouse models of Huntington’s disease [ 20 ]; and Traf6 (TNF receptor-associated factor 6) is an immune response regulator with an E3 ubiquitin ligase activity that has been shown to interact with Sod1 in rat models of amyotrophic lateral sclerosis (ALS), promoting aggregation [ 21 ].…”

Glycine neurotransmission: Its role in development

“…In the adulthood, glycinergic neurotransmission is limited to the spinal cord, retina and few brain areas; however, functional GlyRs have been found almost everywhere in the developing brain. Moreover a variety of evidence strongly support a relevant role of glycine signaling during development; even more, alterations in this signaling has been associated to pathologies in the adulthood (Eichler et al, 2008;Bode and Lynch, 2013;Piton et al, 2013;Pilorge et al, 2016;Morelli et al, 2017;Chen et al, 2022) supporting glycinergic function in proliferation and cell specification and circuit formation. Though, few studies have been carried out to analyze the mechanisms involved.…”

“…Furthermore, recently Chen et al ( 2022) using a combination of molecular modeling and electrophysiology recordings for four novel missense variants in GLRA2 associated with autism spectrum disorder (ASD), identified GLRA2 as the cause of autism spectrum neurodevelopmental phenotypes. The missense variants cause either loss, gain or altered function of GlyR α2 subunit, enlightening the clinical forms associated with human ASD (Chen et al, 2022).…”

Four families with X-linked intellectual disability affected males: Novel deleterious variants and clinical features with the review of literature

“…Recent investigations have noted that missense variants in this gene can result in a loss, gain, or altered function of the encoded protein. In turn, missense variants are likely to either negatively or positively deregulate cortical progenitor homeostasis and neuronal migration in the developing brain, leading to changes in cognition, learning, and memory [21]. The most associated disorder with GLRA2 is X-Linked Intellectual Developmental Disorder, Syndromic, Pilorge Type (MRXSP, OMIM: 301076).…”