Loss of Frrs1l disrupts synaptic AMPA receptor function, and results in neurodevelopmental, motor, cognitive and electrographical abnormalities

Abstract

statementIn this study, we show that the loss of the epilepsy-related gene Frrs1l in mice causes a dramatic reduction in AMPA receptor levels at the synapse. This change elicits severe motor and coordination disabilities, hyperactivity, cognitive defects, behavioural seizures and abnormal electroencephalographic (EEG) patterns. 2 AbstractLoss of function mutations in the human AMPA receptor-associated protein, ferric chelate reductase 1-like (FRRS1L), are associated with a devastating neurological condition incorporating choreoathetosis, cognitive deficits and epileptic encephalopathies. Furthermore, evidence from overexpression and ex vivo studies have implicated FRRS1L in AMPA receptor biogenesis and assembly, suggesting that changes in glutamatergic signalling might underlie the disorder. Here, we investigated the neurological and neurobehavioural correlates of the disorder using a mouse Frrs1l null mutant. The study revealed several neurological defects that mirrored those seen in human patients. We established that mice lacking Frrs1l suffered from a broad spectrum of early-onset motor deficits with no progressive, age-related deterioration. Moreover, Frrs1l -/mice were hyperactive irrespective of test environment, exhibited working memory deficits and displayed significant sleep fragmentation. Longitudinal electroencephalographic recordings also revealed abnormal EEG in Frrs1l -/mice. Parallel investigations into disease aetiology identified a specific deficiency in AMPA receptor levels in the brain of Frrs1l -/mice, while the general levels of several other synaptic components remained unchanged with no obvious alterations in the number of synapses. Furthermore, we established that Frrsl1 deletion results in glycosylation deficits in GLUA2 and GLUA4 AMPA receptor proteins, leading to cytoplasmic retention and a reduction of those specific AMPA receptor levels in the postsynaptic membrane. Overall, this study determines, for the first time in vivo, how loss of FRRS1L function can affect glutamatergic signalling and provides mechanistic insight into the development and progression of a human hyperkinetic disorder.

An ER Assembly Line of AMPA-Receptors Controls Excitatory Neurotransmission and Its Plasticity

“…In this sense, it is noteworthy that FRRS1l expression is strongly upregulated during rodent postnatal development and that impairment of brain function(s) in humans with homozygous mutations of FRRS1l only started after the neonatal period, when children are expected to complete developmental milestones and acquire skills (Brechet et al, 2017). Results similar to some of our own have been reported in independent work on FRRS1lÀ/À animals (Stewart et al, 2019).…”

“…In addition, full proteomic analysis revealed several profound alterations in the AMPAR interactome induced by the homozygous knockout: (1) the binding of CPT1c to GluAs was abolished in line with common binding of FRRS1l and CPT1c to GluAs (Figures 1 and 2); (2) the majority of surface constituents, including GluAs, TARPs 3, 5, 7, and 8, CNIHs, GSG1l, Shisa 6 and 9, PRRT1, or LRRT4, were decreased by 30%-80% ( Figures 3B and 3C) and TARPs 2 and 4 appeared unaltered or mildly reduced; (3) the ER constituents ABHDs 6 and 12 and PORCN were increased, reflecting the additional immature GluA assemblies seen in the ER fraction ( Figure 1E); and (4) the smaller reduction determined for GluAs2-4 compared to GluA1 resulted from their increased amounts in the ER ( Figure S5). Remarkably, all changes in the AMPAR interactome were restricted to the protein level, as steady-state mRNA levels encoding the respective constituents remained unaltered ( Figure S6; see also Stewart et al, 2019).…”

ORANGE: A CRISPR/Cas9-based genome editing toolbox for epitope tagging of endogenous proteins in neurons

“…For instance, we developed knock-in constructs for two AMPA receptor complex constituents, C9orf4 and GSG1L, that have only recently been discovered in a high-resolution proteomics study (53). For both proteins functional characterization is available (54,55), but high-resolution information on subcellular distribution is still lacking due to unavailability of efficient antibodies.…”

Aberrant synaptic release underlies sleep/wake transition deficits in a mouseVamp2mutant

“…Open field behaviour: Mice were placed into one corner of a walled arena (45cm X 45cm) and allowed to explore on two consecutive days for 30 minutes (30). Animal movements and position were tracked using EthoVision XT analysis software (Noldus).…”

“…Grip strength: Grip strength was assessed using a Grip Strength Meter (BioSeb, Chaville, France). Readings were taken from all four paws, three times per mouse as per manufacturer's instructions (30). Measures were averaged and normalised to body weight.…”