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Meltrin beta/ADAM19 interacting with EphA4 in developing neural cells participates in formation of the neuromuscular junction.

Yumoto N, Wakatsuki S, Kurisaki T, Hara Y, Osumi N, Frisén J, Sehara-Fujisawa A - PLoS ONE (2008)

Bottom Line: Meltrin beta plays a regulatory role in formation of the NMJ.The endocytosis of ephrin-Eph complexes is required for efficient contact-dependent repulsion between ephrin and Eph.We propose that Meltrin beta stabilizes the interaction between ephrin-A5 and EphA4 by regulating endocytosis of the ephrinA5-EphA complex negatively, which would contribute to the fine-tuning of the NMJ during development.

View Article: PubMed Central - PubMed

Affiliation: Department of Growth Regulation, Institute for Frontier Medical Sciences, Kyoto University, Shogo-in, Kyoto, Japan.

ABSTRACT

Background: Development of the neuromuscular junction (NMJ) is initiated by the formation of postsynaptic specializations in the central zones of muscles, followed by the arrival of motor nerve terminals opposite the postsynaptic regions. The post- and presynaptic components are then stabilized and modified to form mature synapses. Roles of ADAM (A Disintegrin And Metalloprotease) family proteins in the formation of the NMJ have not been reported previously.

Principal findings: We report here that Meltrin beta, ADAM19, participates in the formation of the NMJ. The zone of acetylcholine receptor alpha mRNA distribution was broader and excess sprouting of motor nerve terminals was more prominent in meltrin beta-deficient than in wild-type embryonic diaphragms. A microarray analysis revealed that the preferential distribution of ephrin-A5 mRNA in the synaptic region of muscles was aberrant in the meltrin beta-deficient muscles. Excess sprouting of motor nerve terminals was also found in ephrin-A5 knockout mice, which lead us to investigate a possible link between Meltrin beta and ephrin-A5-Eph signaling in the development of the NMJ. Meltrin beta and EphA4 interacted with each other in developing motor neurons, and both of these proteins localized in the NMJ. Coexpression of Meltrin beta and EphA4 strongly blocked vesicular internalization of ephrin-A5-EphA4 complexes without requiring the protease activity of Meltrin beta, suggesting a regulatory role of Meltrin beta in ephrin-A5-Eph signaling.

Conclusion: Meltrin beta plays a regulatory role in formation of the NMJ. The endocytosis of ephrin-Eph complexes is required for efficient contact-dependent repulsion between ephrin and Eph. We propose that Meltrin beta stabilizes the interaction between ephrin-A5 and EphA4 by regulating endocytosis of the ephrinA5-EphA complex negatively, which would contribute to the fine-tuning of the NMJ during development.

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Ephrin-A5 is expressed at the NMJ during development and participates in the formation of the NMJ.(A–C) E18.5 intercostal muscles were stained with ephrin-A5 (A) and BTX (B). The BTX-positive postsynaptic apparatus colocalized with ephrin-A5 (C, arrowheads). (D–I) Diaphragm muscles from wild-type (D–F) and ephrin-A5−/− (G–I) mouse embryos at E18.5 were stained with anti-neurofilament (NF) 145 and anti-synaptophysin (SYN) antibodies (D and G) and BTX (E and H). Excess sprouting of the nerve terminals was prominent in the ephrin-A5−/− muscle (I, arrowheads. Compare with the wild-type muscle in F). Arrow: sensory neuron from the lateral part of the diaphragm. Bar: 100 µm.
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pone-0003322-g003: Ephrin-A5 is expressed at the NMJ during development and participates in the formation of the NMJ.(A–C) E18.5 intercostal muscles were stained with ephrin-A5 (A) and BTX (B). The BTX-positive postsynaptic apparatus colocalized with ephrin-A5 (C, arrowheads). (D–I) Diaphragm muscles from wild-type (D–F) and ephrin-A5−/− (G–I) mouse embryos at E18.5 were stained with anti-neurofilament (NF) 145 and anti-synaptophysin (SYN) antibodies (D and G) and BTX (E and H). Excess sprouting of the nerve terminals was prominent in the ephrin-A5−/− muscle (I, arrowheads. Compare with the wild-type muscle in F). Arrow: sensory neuron from the lateral part of the diaphragm. Bar: 100 µm.

Mentions: Previously, Feng et al. showed that motor axons formed topographic maps on muscles and that ephrin-A5 and –A2 were involved in that topographic mapping [26] . They showed that ephrin-A5 was expressed in developing muscles. We investigated the localization of ephrin-A5 by using immunohistochemistry of E18.5 intercostal muscles (Fig. 3A–C). Ephrin-A5 protein was localized at the NMJ (Fig. 3A, C, arrowheads), a distribution that emerged gradually during the late stage of embryogenesis (Figure S1). Then, we performed whole-mount immunostaining of diaphragms from wild-type and ephrin-A5 knockout (ephrin-A5−/−) mice (Fig. 3D–I). In wild-type diaphragms, AChR clusters were tightly expressed in the central endplate zone opposite the nerve terminals (Fig. 3D–F). In contrast, in ephrin-A5−/− mice the axon terminals branched excessively (Fig. 3G–I, arrowheads). Thus, ephrin-A5 also plays a role in the precise positioning of axon terminals apposed to the postsynapses as Meltrin β does.


Meltrin beta/ADAM19 interacting with EphA4 in developing neural cells participates in formation of the neuromuscular junction.

Yumoto N, Wakatsuki S, Kurisaki T, Hara Y, Osumi N, Frisén J, Sehara-Fujisawa A - PLoS ONE (2008)

Ephrin-A5 is expressed at the NMJ during development and participates in the formation of the NMJ.(A–C) E18.5 intercostal muscles were stained with ephrin-A5 (A) and BTX (B). The BTX-positive postsynaptic apparatus colocalized with ephrin-A5 (C, arrowheads). (D–I) Diaphragm muscles from wild-type (D–F) and ephrin-A5−/− (G–I) mouse embryos at E18.5 were stained with anti-neurofilament (NF) 145 and anti-synaptophysin (SYN) antibodies (D and G) and BTX (E and H). Excess sprouting of the nerve terminals was prominent in the ephrin-A5−/− muscle (I, arrowheads. Compare with the wild-type muscle in F). Arrow: sensory neuron from the lateral part of the diaphragm. Bar: 100 µm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2552171&req=5

pone-0003322-g003: Ephrin-A5 is expressed at the NMJ during development and participates in the formation of the NMJ.(A–C) E18.5 intercostal muscles were stained with ephrin-A5 (A) and BTX (B). The BTX-positive postsynaptic apparatus colocalized with ephrin-A5 (C, arrowheads). (D–I) Diaphragm muscles from wild-type (D–F) and ephrin-A5−/− (G–I) mouse embryos at E18.5 were stained with anti-neurofilament (NF) 145 and anti-synaptophysin (SYN) antibodies (D and G) and BTX (E and H). Excess sprouting of the nerve terminals was prominent in the ephrin-A5−/− muscle (I, arrowheads. Compare with the wild-type muscle in F). Arrow: sensory neuron from the lateral part of the diaphragm. Bar: 100 µm.
Mentions: Previously, Feng et al. showed that motor axons formed topographic maps on muscles and that ephrin-A5 and –A2 were involved in that topographic mapping [26] . They showed that ephrin-A5 was expressed in developing muscles. We investigated the localization of ephrin-A5 by using immunohistochemistry of E18.5 intercostal muscles (Fig. 3A–C). Ephrin-A5 protein was localized at the NMJ (Fig. 3A, C, arrowheads), a distribution that emerged gradually during the late stage of embryogenesis (Figure S1). Then, we performed whole-mount immunostaining of diaphragms from wild-type and ephrin-A5 knockout (ephrin-A5−/−) mice (Fig. 3D–I). In wild-type diaphragms, AChR clusters were tightly expressed in the central endplate zone opposite the nerve terminals (Fig. 3D–F). In contrast, in ephrin-A5−/− mice the axon terminals branched excessively (Fig. 3G–I, arrowheads). Thus, ephrin-A5 also plays a role in the precise positioning of axon terminals apposed to the postsynapses as Meltrin β does.

Bottom Line: Meltrin beta plays a regulatory role in formation of the NMJ.The endocytosis of ephrin-Eph complexes is required for efficient contact-dependent repulsion between ephrin and Eph.We propose that Meltrin beta stabilizes the interaction between ephrin-A5 and EphA4 by regulating endocytosis of the ephrinA5-EphA complex negatively, which would contribute to the fine-tuning of the NMJ during development.

View Article: PubMed Central - PubMed

Affiliation: Department of Growth Regulation, Institute for Frontier Medical Sciences, Kyoto University, Shogo-in, Kyoto, Japan.

ABSTRACT

Background: Development of the neuromuscular junction (NMJ) is initiated by the formation of postsynaptic specializations in the central zones of muscles, followed by the arrival of motor nerve terminals opposite the postsynaptic regions. The post- and presynaptic components are then stabilized and modified to form mature synapses. Roles of ADAM (A Disintegrin And Metalloprotease) family proteins in the formation of the NMJ have not been reported previously.

Principal findings: We report here that Meltrin beta, ADAM19, participates in the formation of the NMJ. The zone of acetylcholine receptor alpha mRNA distribution was broader and excess sprouting of motor nerve terminals was more prominent in meltrin beta-deficient than in wild-type embryonic diaphragms. A microarray analysis revealed that the preferential distribution of ephrin-A5 mRNA in the synaptic region of muscles was aberrant in the meltrin beta-deficient muscles. Excess sprouting of motor nerve terminals was also found in ephrin-A5 knockout mice, which lead us to investigate a possible link between Meltrin beta and ephrin-A5-Eph signaling in the development of the NMJ. Meltrin beta and EphA4 interacted with each other in developing motor neurons, and both of these proteins localized in the NMJ. Coexpression of Meltrin beta and EphA4 strongly blocked vesicular internalization of ephrin-A5-EphA4 complexes without requiring the protease activity of Meltrin beta, suggesting a regulatory role of Meltrin beta in ephrin-A5-Eph signaling.

Conclusion: Meltrin beta plays a regulatory role in formation of the NMJ. The endocytosis of ephrin-Eph complexes is required for efficient contact-dependent repulsion between ephrin and Eph. We propose that Meltrin beta stabilizes the interaction between ephrin-A5 and EphA4 by regulating endocytosis of the ephrinA5-EphA complex negatively, which would contribute to the fine-tuning of the NMJ during development.

Show MeSH
Related in: MedlinePlus