PRRT2 Mutant Leads to Dysfunction of Glutamate Signaling

Abstract

Paroxysmal kinesigenic choreoathetosis (PKC) is an inherited disease of the nervous system. We previously identified PRRT2 as the causative gene of PKC. However, as little is known about the function of PRRT2, elucidating its function will benefit not only PKC studies, but also many other related disorders. Here, we reveal higher levels of glutamate in the plasma of PKC patients and the culture medium of neurons following knock-out Prrt2 expression. Using double immunostaining assays we confirm Prrt2 is located at the glutamatergic neurons in accordance with its function. Our co-immunoprecipitation assays reveal mutant PRRT2 interferes with SNAP25 and GRIA1 interactions, respectively. Furthermore, using live-labeling techniques, we confirmed co-transfection with mutant PRRT2 caused an increase in GRIA1 distribution on the cell surface. Therefore, our results suggest that mutant PRRT2, probably through its weakened interaction with SNAP25, affects glutamate signaling and glutamate receptor activity, resulting in the increase of glutamate release and subsequent neuronal hyperexcitability.

PRRT2 mutations lead to neuronal dysfunction and neurodevelopmental defects

“…We also examined the distribution of EGFP-Prrt2 in transfected cultured HEK293T cells (Figure 2B, right panel). The fluorescence signals from EGFP and Prrt2 antibody predominantly co-localized at the cell membrane, consistent with previous observations using other protein tags [1, 20]. To further test the specificity of the antibody, we test whether knocking down Prrt2 expression in cells by RNA interference (RNAi) will eliminate antibody reactivity (Figure 2C).…”

“…Recent studies have revealed that PRRT2 is a component of the AMPAR (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor) complex [30, 31] and may be involved in neurotransmitter release and glutamate signaling[20, 32]. Consistently, PRRT2 was found to co-localize with pre- and post-synaptic markers vGlut1 and PSD95 in our study.…”

The epileptic and nonepileptic spectrum of paroxysmal dyskinesias: Channelopathies, synaptopathies, and transportopathies

“…There is evidence that PRRT2 localizes mostly in axons of glutamatergic synapses [41,42]. However, previous works have suggested that it interacts with the SNARE protein SNAP-25 as well as the GluA1 subunit of the AMPA-type glutamate receptor complex, leaving the question open of whether PRRT2 localizes at the presynaptic terminal, postsynaptic terminal, or both.…”

“…Finally, proteomic studies demonstrated an interaction of PRRT2 with the AMPA receptor complex [49]. After infection with shRNA-Prrt2 lentivirus, there is an increase in glutamate levels in the culture medium of neurons as well as in the distribution of GluA1 on the cell membrane [42]. One of the reasons why carbamazepine is very effective in patients with PRRT2 mutations might be owing to its ability to enhance the activity of the glutamate transporter 3 at the post-synaptic terminal [50], thus allowing glutamate to be pumped into postsynaptic terminals without activating its receptor.…”

A homozygous PIGN missense mutation in Soft-Coated Wheaten Terriers with a canine paroxysmal dyskinesia

“…A minority of patients with heterozygous SLC2A1 mutations develop isolated PED [11, 15–17]. PRRT2 encodes proline-rich transmembrane protein 2, which interacts with SNAP25 and thereby influences the release of glutamate or other neurotransmitters [18]. Heterozygous PRRT2 mutations have been found in patients with several paroxysmal neurologic disorders including PKD [19].…”