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Juxtaparanodal clustering of Shaker-like K+ channels in myelinated axons depends on Caspr2 and TAG-1.

Poliak S, Salomon D, Elhanany H, Sabanay H, Kiernan B, Pevny L, Stewart CL, Xu X, Chiu SY, Shrager P, Furley AJ, Peles E - J. Cell Biol. (2003)

Bottom Line: In myelinated axons, K+ channels are concealed under the myelin sheath in the juxtaparanodal region, where they are associated with Caspr2, a member of the neurexin superfamily.Deletion of Caspr2 in mice by gene targeting revealed that it is required to maintain K+ channels at this location.These results demonstrate that Caspr2 and TAG-1 form a scaffold that is necessary to maintain K+ channels at the juxtaparanodal region, suggesting that axon-glia interactions mediated by these proteins allow myelinating glial cells to organize ion channels in the underlying axonal membrane.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot 76100, Israel.

ABSTRACT
In myelinated axons, K+ channels are concealed under the myelin sheath in the juxtaparanodal region, where they are associated with Caspr2, a member of the neurexin superfamily. Deletion of Caspr2 in mice by gene targeting revealed that it is required to maintain K+ channels at this location. Furthermore, we show that the localization of Caspr2 and clustering of K+ channels at the juxtaparanodal region depends on the presence of TAG-1, an immunoglobulin-like cell adhesion molecule that binds Caspr2. These results demonstrate that Caspr2 and TAG-1 form a scaffold that is necessary to maintain K+ channels at the juxtaparanodal region, suggesting that axon-glia interactions mediated by these proteins allow myelinating glial cells to organize ion channels in the underlying axonal membrane.

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Reduced juxtaparanodal accumulation of K+ channels in the PNS of Caspr2−/− mice. (A–F) Images showing immunofluorescence staining of sciatic nerve sections from wild-type (A–C) or Caspr2-deficient mice (D–F), using antibodies to Kv1.2 (red; A and D) and Caspr (green; B and E) as indicated. Merge images are shown on the right (C and F). Representative images of the nodal region from teased fiber preparations are shown in the insets in C and F. (G and H) Teased sciatic nerves from wild-type (G) and Caspr2- (H) mice, labeled with antibodies to Kv1.2 (green) and Caspr (red). (I) Quantification of the percentage of juxtaparanodes exhibiting normal appearance of Kv1.2, identified as the domain adjacent to Caspr-stained area (n = +/+ 174, −/− 219). (J) Immunolabeling of teased sciatic nerve from Caspr2−/− mutant, showing intense staining of Kv1.2 along the internodes, entering the juxtaparanodal region. (Inset) Western blot showing the expression of Kv1.2 in sciatic nerve lysates from wild-type (+/+) and Caspr2- (−/−) mice. (K) Higher magnification of the labeled frame in J. (L) Another example of the localization of K+ channels in the mutant, present in a double line crossing the juxtaparanodal region and terminating as a ring at the border between the juxtaparanodes and paranodes. The location of the paranodes is marked (P) in K and L. Bars: A–H, 20 μm; J–L, 10 μm of the bar shown in J.
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fig4: Reduced juxtaparanodal accumulation of K+ channels in the PNS of Caspr2−/− mice. (A–F) Images showing immunofluorescence staining of sciatic nerve sections from wild-type (A–C) or Caspr2-deficient mice (D–F), using antibodies to Kv1.2 (red; A and D) and Caspr (green; B and E) as indicated. Merge images are shown on the right (C and F). Representative images of the nodal region from teased fiber preparations are shown in the insets in C and F. (G and H) Teased sciatic nerves from wild-type (G) and Caspr2- (H) mice, labeled with antibodies to Kv1.2 (green) and Caspr (red). (I) Quantification of the percentage of juxtaparanodes exhibiting normal appearance of Kv1.2, identified as the domain adjacent to Caspr-stained area (n = +/+ 174, −/− 219). (J) Immunolabeling of teased sciatic nerve from Caspr2−/− mutant, showing intense staining of Kv1.2 along the internodes, entering the juxtaparanodal region. (Inset) Western blot showing the expression of Kv1.2 in sciatic nerve lysates from wild-type (+/+) and Caspr2- (−/−) mice. (K) Higher magnification of the labeled frame in J. (L) Another example of the localization of K+ channels in the mutant, present in a double line crossing the juxtaparanodal region and terminating as a ring at the border between the juxtaparanodes and paranodes. The location of the paranodes is marked (P) in K and L. Bars: A–H, 20 μm; J–L, 10 μm of the bar shown in J.

Mentions: Next, we examined the distribution of K+ channels in sciatic nerve of Caspr2−/− mice. Double immunofluorescence using antibodies to Caspr and Kv1.2 revealed a similar marked reduction in the accumulation of K+ channels at the juxtaparanodal region in the PNS as observed in the CNS (Fig. 4, A–F). The same results were obtained using antibodies to KV1.1 and Kvβ2 subunits in either frozen sections or teased fiber preparations. Although strong juxtaparanodal accumulation of K+ channels was recorded in 90.3 ± 3.1% of the sites in sciatic nerve sections from wild-type mice, it was only detected in 10.4 ± 6.7% of these sites in the mutant (Fig. 4 I). In most cases, juxtaparanodal labeling of K+ channels in the mutant was detected near the paranodes, which occasionally overlapped with the edge of Caspr staining (Fig. 4 H). In peripheral nerves of wild-type mice, K+ channels flank Caspr in a double strand that apposes the inner mesaxon of the myelin sheath throughout the internodes (the juxtamesaxon; Arroyo et al., 1999; Poliak et al., 2001). In contrast to the reduced accumulation of K+ channels in the juxtaparanodal region, internodal localization of K+ channels appeared normal in the sciatic nerve of Caspr2−/− (Fig. 4, H and J), suggesting that different mechanisms control the localization of K+ channels in the juxtaparanodes and along the internodes. Notably, the double juxtamesaxonal lines of K+ channels extended into the juxtaparanodal region and ended as a ring at the border of the paranodes (Fig. 4, J–L). Western blot of sciatic nerve lysates using an antibody to Kv1.2 (Fig. 4 J, inset), showed that similar amounts of K+ channels are found in Caspr2 mutant and wild-type nerves, indicating that the reduction of these channels at the juxtaparanodal region resulted from their redistribution along the internodes. Altogether, these results demonstrate that Caspr2 is required for the localization of K+ channels at the juxtaparanodal region in both CNS and PNS myelinated axons.


Juxtaparanodal clustering of Shaker-like K+ channels in myelinated axons depends on Caspr2 and TAG-1.

Poliak S, Salomon D, Elhanany H, Sabanay H, Kiernan B, Pevny L, Stewart CL, Xu X, Chiu SY, Shrager P, Furley AJ, Peles E - J. Cell Biol. (2003)

Reduced juxtaparanodal accumulation of K+ channels in the PNS of Caspr2−/− mice. (A–F) Images showing immunofluorescence staining of sciatic nerve sections from wild-type (A–C) or Caspr2-deficient mice (D–F), using antibodies to Kv1.2 (red; A and D) and Caspr (green; B and E) as indicated. Merge images are shown on the right (C and F). Representative images of the nodal region from teased fiber preparations are shown in the insets in C and F. (G and H) Teased sciatic nerves from wild-type (G) and Caspr2- (H) mice, labeled with antibodies to Kv1.2 (green) and Caspr (red). (I) Quantification of the percentage of juxtaparanodes exhibiting normal appearance of Kv1.2, identified as the domain adjacent to Caspr-stained area (n = +/+ 174, −/− 219). (J) Immunolabeling of teased sciatic nerve from Caspr2−/− mutant, showing intense staining of Kv1.2 along the internodes, entering the juxtaparanodal region. (Inset) Western blot showing the expression of Kv1.2 in sciatic nerve lysates from wild-type (+/+) and Caspr2- (−/−) mice. (K) Higher magnification of the labeled frame in J. (L) Another example of the localization of K+ channels in the mutant, present in a double line crossing the juxtaparanodal region and terminating as a ring at the border between the juxtaparanodes and paranodes. The location of the paranodes is marked (P) in K and L. Bars: A–H, 20 μm; J–L, 10 μm of the bar shown in J.
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fig4: Reduced juxtaparanodal accumulation of K+ channels in the PNS of Caspr2−/− mice. (A–F) Images showing immunofluorescence staining of sciatic nerve sections from wild-type (A–C) or Caspr2-deficient mice (D–F), using antibodies to Kv1.2 (red; A and D) and Caspr (green; B and E) as indicated. Merge images are shown on the right (C and F). Representative images of the nodal region from teased fiber preparations are shown in the insets in C and F. (G and H) Teased sciatic nerves from wild-type (G) and Caspr2- (H) mice, labeled with antibodies to Kv1.2 (green) and Caspr (red). (I) Quantification of the percentage of juxtaparanodes exhibiting normal appearance of Kv1.2, identified as the domain adjacent to Caspr-stained area (n = +/+ 174, −/− 219). (J) Immunolabeling of teased sciatic nerve from Caspr2−/− mutant, showing intense staining of Kv1.2 along the internodes, entering the juxtaparanodal region. (Inset) Western blot showing the expression of Kv1.2 in sciatic nerve lysates from wild-type (+/+) and Caspr2- (−/−) mice. (K) Higher magnification of the labeled frame in J. (L) Another example of the localization of K+ channels in the mutant, present in a double line crossing the juxtaparanodal region and terminating as a ring at the border between the juxtaparanodes and paranodes. The location of the paranodes is marked (P) in K and L. Bars: A–H, 20 μm; J–L, 10 μm of the bar shown in J.
Mentions: Next, we examined the distribution of K+ channels in sciatic nerve of Caspr2−/− mice. Double immunofluorescence using antibodies to Caspr and Kv1.2 revealed a similar marked reduction in the accumulation of K+ channels at the juxtaparanodal region in the PNS as observed in the CNS (Fig. 4, A–F). The same results were obtained using antibodies to KV1.1 and Kvβ2 subunits in either frozen sections or teased fiber preparations. Although strong juxtaparanodal accumulation of K+ channels was recorded in 90.3 ± 3.1% of the sites in sciatic nerve sections from wild-type mice, it was only detected in 10.4 ± 6.7% of these sites in the mutant (Fig. 4 I). In most cases, juxtaparanodal labeling of K+ channels in the mutant was detected near the paranodes, which occasionally overlapped with the edge of Caspr staining (Fig. 4 H). In peripheral nerves of wild-type mice, K+ channels flank Caspr in a double strand that apposes the inner mesaxon of the myelin sheath throughout the internodes (the juxtamesaxon; Arroyo et al., 1999; Poliak et al., 2001). In contrast to the reduced accumulation of K+ channels in the juxtaparanodal region, internodal localization of K+ channels appeared normal in the sciatic nerve of Caspr2−/− (Fig. 4, H and J), suggesting that different mechanisms control the localization of K+ channels in the juxtaparanodes and along the internodes. Notably, the double juxtamesaxonal lines of K+ channels extended into the juxtaparanodal region and ended as a ring at the border of the paranodes (Fig. 4, J–L). Western blot of sciatic nerve lysates using an antibody to Kv1.2 (Fig. 4 J, inset), showed that similar amounts of K+ channels are found in Caspr2 mutant and wild-type nerves, indicating that the reduction of these channels at the juxtaparanodal region resulted from their redistribution along the internodes. Altogether, these results demonstrate that Caspr2 is required for the localization of K+ channels at the juxtaparanodal region in both CNS and PNS myelinated axons.

Bottom Line: In myelinated axons, K+ channels are concealed under the myelin sheath in the juxtaparanodal region, where they are associated with Caspr2, a member of the neurexin superfamily.Deletion of Caspr2 in mice by gene targeting revealed that it is required to maintain K+ channels at this location.These results demonstrate that Caspr2 and TAG-1 form a scaffold that is necessary to maintain K+ channels at the juxtaparanodal region, suggesting that axon-glia interactions mediated by these proteins allow myelinating glial cells to organize ion channels in the underlying axonal membrane.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Cell Biology, The Weizmann Institute of Science, Rehovot 76100, Israel.

ABSTRACT
In myelinated axons, K+ channels are concealed under the myelin sheath in the juxtaparanodal region, where they are associated with Caspr2, a member of the neurexin superfamily. Deletion of Caspr2 in mice by gene targeting revealed that it is required to maintain K+ channels at this location. Furthermore, we show that the localization of Caspr2 and clustering of K+ channels at the juxtaparanodal region depends on the presence of TAG-1, an immunoglobulin-like cell adhesion molecule that binds Caspr2. These results demonstrate that Caspr2 and TAG-1 form a scaffold that is necessary to maintain K+ channels at the juxtaparanodal region, suggesting that axon-glia interactions mediated by these proteins allow myelinating glial cells to organize ion channels in the underlying axonal membrane.

Show MeSH
Related in: MedlinePlus