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[Beta]IV-spectrin regulates sodium channel clustering through ankyrin-G at axon initial segments and nodes of Ranvier.

Komada M, Soriano P - J. Cell Biol. (2002)

Bottom Line: In betaIV-spectrin- neurons, neither ankyrin-G nor voltage-gated sodium channels (VGSC) are correctly clustered at these sites, suggesting that impaired action potential caused by mislocalization of VGSC leads to the phenotype.Conversely, in ankyrin-G- neurons, betaIV-spectrin is not localized to these sites.These results indicate that betaIV-spectrin and ankyrin-G mutually stabilize the membrane protein cluster and the linked membrane cytoskeleton at AIS and NR.

View Article: PubMed Central - PubMed

Affiliation: Program in Developmental Biology and Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA. makomada@bio.titech.ac.jp

ABSTRACT
beta-Spectrin and ankyrin are major components of the membrane cytoskeleton. We have generated mice carrying a mutation in the betaIV-spectrin gene using gene trapping in embryonic stem cells. Mice homozygous for the mutation exhibit tremors and contraction of hindlimbs. betaIV-spectrin expression is mostly restricted to neurons, where it colocalizes with and binds to ankyrin-G at axon initial segments (AISs) and nodes of Ranvier (NR). In betaIV-spectrin- neurons, neither ankyrin-G nor voltage-gated sodium channels (VGSC) are correctly clustered at these sites, suggesting that impaired action potential caused by mislocalization of VGSC leads to the phenotype. Conversely, in ankyrin-G- neurons, betaIV-spectrin is not localized to these sites. These results indicate that betaIV-spectrin and ankyrin-G mutually stabilize the membrane protein cluster and the linked membrane cytoskeleton at AIS and NR.

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Clustering of Nav1.6 at NR of βIV-spectrin– sciatic nerves. Double staining of wild-type (A, A′, and A′′) and βIV-spectrin– (B, B′, and B′′) sciatic nerves with anti–βIV-spectrin (A and B) and anti-Nav1.6 (A′ and B′) antibodies. (A′′) and (B′′) are merged images. Arrowheads in A′, B′, A′′, and B′′ indicate some of the Nav1.6-positive NR which are also positive for βIV-spectrin in the wild-type (A). Insets in A′ and B′ show enlarged images of typical NR. In C, the number of Nav1.6-positive NR in wild-type (+/+) and βIV-spectrin– (−/−) sciatic nerve sections were quantified. Mean numbers per 0.1 mm2 field are shown with SD (n = 9 for each genotype). Mice were killed at 3 mo of age. Bars, 100 μm.
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fig7: Clustering of Nav1.6 at NR of βIV-spectrin– sciatic nerves. Double staining of wild-type (A, A′, and A′′) and βIV-spectrin– (B, B′, and B′′) sciatic nerves with anti–βIV-spectrin (A and B) and anti-Nav1.6 (A′ and B′) antibodies. (A′′) and (B′′) are merged images. Arrowheads in A′, B′, A′′, and B′′ indicate some of the Nav1.6-positive NR which are also positive for βIV-spectrin in the wild-type (A). Insets in A′ and B′ show enlarged images of typical NR. In C, the number of Nav1.6-positive NR in wild-type (+/+) and βIV-spectrin– (−/−) sciatic nerve sections were quantified. Mean numbers per 0.1 mm2 field are shown with SD (n = 9 for each genotype). Mice were killed at 3 mo of age. Bars, 100 μm.

Mentions: Next, we examined the localization of Nav1.6 at NR of the peripheral nervous system. In the βIV-spectrin– sciatic nerves, Nav1.6-positive but βIV-spectrin–negative NR were detected (Fig. 7 , B-B′′). However, the number of Nav1.6-positive NR was less than in the wild-type littermate in which the NR were also positive for βIV-spectrin (Fig. 7, A–A′′). The number of Nav1.6-positive NR was quantified in sciatic nerve sections of wild-type and the mutant mice. Counting of nine randomly chosen 0.1-mm2 fields from three mice of each genotype revealed that the numbers were 98 ± 23 and 44 ± 15 (mean ± SD) per field in the wild-type and the mutant, respectively (55% reduction in the mutant; Fig. 7 C). In addition, anti-Nav1.6 staining of the NR was often fainter in the mutant than in the wild-type control (Fig. 7, A′ and B′, insets). Phase contrast microscopy showed that the number as well as the morphology of NR was unaffected in the mutant (unpublished data).


[Beta]IV-spectrin regulates sodium channel clustering through ankyrin-G at axon initial segments and nodes of Ranvier.

Komada M, Soriano P - J. Cell Biol. (2002)

Clustering of Nav1.6 at NR of βIV-spectrin– sciatic nerves. Double staining of wild-type (A, A′, and A′′) and βIV-spectrin– (B, B′, and B′′) sciatic nerves with anti–βIV-spectrin (A and B) and anti-Nav1.6 (A′ and B′) antibodies. (A′′) and (B′′) are merged images. Arrowheads in A′, B′, A′′, and B′′ indicate some of the Nav1.6-positive NR which are also positive for βIV-spectrin in the wild-type (A). Insets in A′ and B′ show enlarged images of typical NR. In C, the number of Nav1.6-positive NR in wild-type (+/+) and βIV-spectrin– (−/−) sciatic nerve sections were quantified. Mean numbers per 0.1 mm2 field are shown with SD (n = 9 for each genotype). Mice were killed at 3 mo of age. Bars, 100 μm.
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fig7: Clustering of Nav1.6 at NR of βIV-spectrin– sciatic nerves. Double staining of wild-type (A, A′, and A′′) and βIV-spectrin– (B, B′, and B′′) sciatic nerves with anti–βIV-spectrin (A and B) and anti-Nav1.6 (A′ and B′) antibodies. (A′′) and (B′′) are merged images. Arrowheads in A′, B′, A′′, and B′′ indicate some of the Nav1.6-positive NR which are also positive for βIV-spectrin in the wild-type (A). Insets in A′ and B′ show enlarged images of typical NR. In C, the number of Nav1.6-positive NR in wild-type (+/+) and βIV-spectrin– (−/−) sciatic nerve sections were quantified. Mean numbers per 0.1 mm2 field are shown with SD (n = 9 for each genotype). Mice were killed at 3 mo of age. Bars, 100 μm.
Mentions: Next, we examined the localization of Nav1.6 at NR of the peripheral nervous system. In the βIV-spectrin– sciatic nerves, Nav1.6-positive but βIV-spectrin–negative NR were detected (Fig. 7 , B-B′′). However, the number of Nav1.6-positive NR was less than in the wild-type littermate in which the NR were also positive for βIV-spectrin (Fig. 7, A–A′′). The number of Nav1.6-positive NR was quantified in sciatic nerve sections of wild-type and the mutant mice. Counting of nine randomly chosen 0.1-mm2 fields from three mice of each genotype revealed that the numbers were 98 ± 23 and 44 ± 15 (mean ± SD) per field in the wild-type and the mutant, respectively (55% reduction in the mutant; Fig. 7 C). In addition, anti-Nav1.6 staining of the NR was often fainter in the mutant than in the wild-type control (Fig. 7, A′ and B′, insets). Phase contrast microscopy showed that the number as well as the morphology of NR was unaffected in the mutant (unpublished data).

Bottom Line: In betaIV-spectrin- neurons, neither ankyrin-G nor voltage-gated sodium channels (VGSC) are correctly clustered at these sites, suggesting that impaired action potential caused by mislocalization of VGSC leads to the phenotype.Conversely, in ankyrin-G- neurons, betaIV-spectrin is not localized to these sites.These results indicate that betaIV-spectrin and ankyrin-G mutually stabilize the membrane protein cluster and the linked membrane cytoskeleton at AIS and NR.

View Article: PubMed Central - PubMed

Affiliation: Program in Developmental Biology and Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA. makomada@bio.titech.ac.jp

ABSTRACT
beta-Spectrin and ankyrin are major components of the membrane cytoskeleton. We have generated mice carrying a mutation in the betaIV-spectrin gene using gene trapping in embryonic stem cells. Mice homozygous for the mutation exhibit tremors and contraction of hindlimbs. betaIV-spectrin expression is mostly restricted to neurons, where it colocalizes with and binds to ankyrin-G at axon initial segments (AISs) and nodes of Ranvier (NR). In betaIV-spectrin- neurons, neither ankyrin-G nor voltage-gated sodium channels (VGSC) are correctly clustered at these sites, suggesting that impaired action potential caused by mislocalization of VGSC leads to the phenotype. Conversely, in ankyrin-G- neurons, betaIV-spectrin is not localized to these sites. These results indicate that betaIV-spectrin and ankyrin-G mutually stabilize the membrane protein cluster and the linked membrane cytoskeleton at AIS and NR.

Show MeSH
Related in: MedlinePlus