A hot spot for hotfoot mutations in the gene encoding the δ2 glutamate receptor

Abstract

The orphan glutamate receptor delta2 is selectively expressed in Purkinje cells and plays a crucial role in cerebellar functions. Recently, ataxia in the hotfoot mouse ho4J was demonstrated to be caused by a deletion in the delta2 receptor gene (Grid2) removing the N-terminal 170 amino acids of the delta2 receptor. To understand how delta2 receptors function, we characterized mutations in eight additional spontaneously occurring hotfoot alleles of Grid2. The mouse Grid2 gene consists of 16 exons, spanning approximately 1.4 Mb. Genomic DNA analysis showed that seven hotfoot mutants had a deletion of one or more exons encoding the N-terminal domain of delta2 receptors. The exception is ho5J, which has a point mutation in exon 12. Deletions in ho7J, ho9J, ho11J and ho12J mice result in the in-frame deletion of between 40 and 95 amino acids. Expression of constructs containing these deletions in HEK293 cells resulted in protein retention in the endoplasmic reticulum or cis-Golgi without transport to the cell surface. Coimmunoprecipitation assays indicated that these deletions also reduce the intermolecular interaction between individual delta2 receptors. These results indicate that the deleted N-terminal regions are crucial for oligomerization of delta2 receptors and their subsequent transport to the cell surface of Purkinje cells. The relatively large size of the Grid2 gene may be one of the reasons why many spontaneous mutations occur in this gene. In addition, the frequent occurrence of in-frame deletions within the N-terminal domain in hotfoot mutants suggests the importance of this domain in the function of delta2 receptors.

Characterization of a conserved aphidicolin-sensitive common fragile site at human 4q22 and mouse 6C1: possible association with an inherited disease and cancer

“…In total, 11 recessive mutations abolishing grid2 expression were analysed at the molecular level. Among them, nine result from deletions of one or more exons and one corresponds to the spontaneous integration of an exogenous DNA sequence (Figure 2b) (Lalouette et al, 2001;Wang et al, 2003). Interestingly, all the mutations lie in the part of grid2 that is especially prone to breakage (Figure 1c, Figure 2d).…”

A New Motif Necessary and Sufficient for Stable Localization of the δ2 Glutamate Receptors at Postsynaptic Spines

“…Interestingly, of the 20 alleles known so far, most mutants retain mutant GluR␦2 in the endoplasmic reticulum, indicating that GluR␦2 must be transported to the Purkinje cell surface for it to function properly (14,15,21). In addition, we recently demonstrated that the application of an antibody against the extracellular domain of GluR␦2 induced endocytosis of the AMPA receptor GluR2 and inhibited further induction of LTD (17).…”

“…In addition, although 50 -70% of the iGluRs are detected in the intracellular compartments of neurons (12,13), GluR␦2 is predominantly expressed at the cell surface (14). Interestingly, in the ataxic mutant mice hotfoot-4J, -7J, -11J, and -12J, GluR␦2 failed to be transported to the cell surface (14,15), which suggests that efficient trafficking of GluR␦2 to the cell surface is essential for its functioning in the cerebellum. Furthermore, GluR␦2 is not only transported to the Purkinje cell surface but also to spine regions where parallel fibers form synapses (16).…”

Recent Developments in Gene Therapy Research Targeted to Cerebellar Disorders

“…Recently, the genes responsible for those mutants have been identified and include missense mutations, nonsense mutations or frameshift mutations in the causative genes. The hotfoot mice are spontaneously occurring recessive mutants (Guastavino et al, 1990) that carry mutations in Grid2, which encodes the 2 glutamate receptor (GluR2) (Wang et al, 2003). To date, more than 10 mutant alleles have been identified; among these, hotfoot5J mice have been shown to carry a point mutation in exon 12 of Grid2, which creates a stop codon in the region encoding transmembrane 3 of GluR2 protein (Wang et al, 2003).…”

“…The hotfoot mice are spontaneously occurring recessive mutants (Guastavino et al, 1990) that carry mutations in Grid2, which encodes the 2 glutamate receptor (GluR2) (Wang et al, 2003). To date, more than 10 mutant alleles have been identified; among these, hotfoot5J mice have been shown to carry a point mutation in exon 12 of Grid2, which creates a stop codon in the region encoding transmembrane 3 of GluR2 protein (Wang et al, 2003). The aberrant GluR2 protein is easily degraded and is not detected in Purkinje cells of hotfoot5J mice; therefore, hotfoot5J mice are mutants lacking GluR2 function.…”