Tyrosine Phosphorylation of Nicotinic Acetylcholine Receptor Mediates Grb2 Binding

Colledge, Marcie; Froehner, Stanley C.

doi:10.1523/jneurosci.17-13-05038.1997

Cited by 45 publications

(31 citation statements)

References 66 publications

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“…This suggests that β subunit phosphorylation regulates protein interactions involved in receptor localization, whereas δ loop phosphorylation serves other functions. Indeed, tyrosine-phosphorylated δ subunit of Torpedo AChR binds the cytoplasmic tyrosine kinases fyn and fyk (Swope and Huganir, 1994), and the adaptor protein, Grb2, via their SH2 domains, consistent with a signaling role (Colledge and Froehner, 1997).…”

Section: Phosphorylation Of Achr β and δ Subunitsmentioning

confidence: 75%

Rapsyn carboxyl terminal domains mediate muscle specific kinase–induced phosphorylation of the muscle acetylcholine receptor

et al. 2008

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At the developing vertebrate neuromuscular junction, postsynaptic localization of the acetylcholine receptor (AChR) is regulated by agrin signaling via the muscle specific kinase (MuSK) and requires an intracellular scaffolding protein called rapsyn. In addition to its structural role, rapsyn is also necessary for agrin-induced tyrosine phosphorylation of the AChR, which regulates some aspects of receptor localization. Here, we have investigated the molecular mechanism by which rapsyn mediates AChR phosphorylation. In a heterologous COS cell system, we show that MuSK and rapsyn induced phosphorylation of β subunit tyrosine 390 (Y390) and δ subunit Y393, as in muscle cells. Mutation of β Y390 or δ Y393 did not inhibit MuSK/rapsyn-induced phosphorylation of the other subunit in COS cells, and mutation of β Y390 did not inhibit agrin-induced phosphorylation of the δ subunit in Sol8 muscle cells; thus, their phosphorylation occurs independently, downstream of MuSK activation. In COS cells, we further show that MuSK-induced phosphorylation of the β subunit was mediated by rapsyn, as MuSK plus rapsyn increased β Y390 phosphorylation more than rapsyn alone and MuSK alone had no effect. Intriguingly, MuSK also induced tyrosine phosphorylation of rapsyn itself. We then used deletion mutants to map the rapsyn domains responsible for activation of cytoplasmic tyrosine kinases that phosphorylate the AChR subunits. We found that rapsyn C-terminal domains (amino acids 212-412) are both necessary and sufficient for activation of tyrosine kinases and induction of cellular tyrosine phosphorylation. Moreover, deletion of the rapsyn RING domain (365-412) abolished MuSK-induced tyrosine phosphorylation of the AChR β subunit. Together, these findings suggest that rapsyn facilitates AChR phosphorylation by activating or localizing tyrosine kinases via its C-terminal domains. Keywords neuromuscular junction; synaptogenesis; agrin; postsynaptic membraneAt the developing neuromuscular junction in vertebrates, several nerve-derived signals combine to localize the acetylcholine receptor at postsynaptic sites (Sanes and Lichtman, 2001, Burden, 2002, Kummer et al., 2006. One essential factor is agrin, which signals via the MuSK receptor tyrosine kinase and induces and/or stabilizes clustering of the AChR in the postsynaptic membrane (reviewed in (Kummer et al., 2006 Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. is prepatterned and AChR clusters occur in the central region of the muscle prior to and even in the absence of neural innervation (Lin et al., 2001, Yang et al., 2001). However, upo...

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Section: Phosphorylation Of Achr β and δ Subunitsmentioning

confidence: 75%

Rapsyn carboxyl terminal domains mediate muscle specific kinase–induced phosphorylation of the muscle acetylcholine receptor

et al. 2008

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“…Results from studies of neuregulin-1 and ErbB2 mutant mice indicate that neuregulin is essential for the formation and maintenance of the neuromuscular junction (43,54). In support of this hypothesis are findings that ErbB protein tyrosine kinases and downstream signaling molecules are concentrated at the neuromuscular junction (3,15,16,40,55,56). However, the mechanisms underlying clustering of ErbB proteins in muscle cells remain unclear.…”

Section: Fig 9 Interaction Between Erbin and Psd-95mentioning

confidence: 99%

Erbin Is a Protein Concentrated at Postsynaptic Membranes That Interacts with PSD-95

Huang

Wang

Xiong³

et al. 2001

Journal of Biological Chemistry

124

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Neuregulin is a factor essential for synapse-specific transcription of acetylcholine receptor genes at the neuromuscular junction. Its receptors, ErbB receptor tyrosine kinases, are localized at the postjunctional membrane presumably to ensure localized signaling. However, the molecular mechanisms underlying synaptic localization of ErbBs are unknown. The neuromuscular junction is a cholinergic synapse that conveys signals rapidly from motoneurons to muscle cells. The fast and accurate neurotransmission at this synapse is guaranteed by the high concentration of acetylcholine receptors in the postsynaptic membrane, which accounts for only 0.1% of total muscle surface (1, 2). Muscle fibers are multinucleated cells. Remarkably, it is only the synaptic nuclei that actively transcribe genes encoding acetylcholine receptor subunits. Such synapse-specific transcription is mediated by neuregulin, a factor used by motoneurons to stimulate acetylcholine receptor synthesis at the neuromuscular junction. Neuregulin receptors are transmembrane tyrosine kinases of the ErbB family: ErbB2, ErbB3, and ErbB4. Stimulation by neuregulin of ErbB proteins leads to their tyrosine phosphorylation (3-6) and subsequent activation of multiple intracellular signaling cascades (6 -9), essential for compartmental synthesis of acetylcholine receptors. In the central nervous system, neuregulin regulates expression of neuronal nicotinic acetylcholine receptor (10), N-methyl-D-aspartate receptor (11), and ␥-aminobutyric acid receptor (12). Recent studies suggest that in addition to an essential role during development, neuregulin appears to regulate synaptic plasticity in the adult brain (13).ErbB proteins are not expressed evenly on the surface of cells. On the contrary, they are localized in subcellular compartments. In the nervous system, ErbB proteins are concentrated in postsynaptic membranes both at the neuromuscular junction (3, 14 -16) and in the central nervous system (13,17). In epithelial cells, ErbB2 appears to be enriched in basolateral membranes (18). The mechanism by which ErbB proteins are localized in the subcellular compartments remains largely unknown. The intracellular portions of ErbB receptor tyrosine kinases contain large C termini in addition to kinase domains. Thus, it is conceivable that ErbBs may interact with proteins that regulate their localization, surface expression, or kinase activity. Indeed, recent studies demonstrated that ErbB4, via its C terminus, interacts with postsynaptic density (PSD) 1 -95 (or SAP90), a PDZ domain-containing protein (13,17). PDZ domains are motifs of 80 -90 amino acids which often bind to specific sequences at the extreme C termini of target proteins (19 -22). They were originally identified in PSD-95, the Drosophila septate junction protein discs large, and the epithelial tight-junction protein zona occludens 1 (23-26). PDZ domaincontaining proteins appear to coordinate the assembly of functional subcellular domains. PSD-95 uses multiple PDZ domains to cluster ion channels, receptors, ...

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“…Nonetheless, the tyrosine phosphorylation sites in the ␤-and ␦-subunits are embedded in different sequences, indicating that the phosphorylated subunits are unlikely to recruit the same adaptor protein (Colledge and Froehner, 1997). Notably, the tyrosine phosphorylation site in the ␦-subunit conforms to a binding site for SH2 domains, whereas the tyrosine phosphorylation site in the ␤-subunit is not predicted to bind SH2 or PTB domains.…”

Section: Research Articlementioning

confidence: 99%

Synaptic differentiation is defective in mice lacking acetylcholine receptor β-subunit tyrosine phosphorylation

Friese¹,

Blagden²,

Burden³

2007

Development

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Agrin activates MuSK, a receptor tyrosine kinase expressed in skeletal muscle, leading to tyrosine phosphorylation of the acetylcholine receptor (AChR) ␤-subunit and clustering of AChRs. The importance of AChR ␤-subunit tyrosine phosphorylation in clustering AChRs and regulating synaptic differentiation is poorly understood. We generated mice with targeted mutations in the three intracellular tyrosines of the AChR ␤-subunit (AChR-␤ 3F/3F). Mice lacking AChR ␤-subunit tyrosine phosphorylation thrive postnatally and have no overt behavioral defects, indicating that AChR ␤-subunit tyrosine phosphorylation is not essential for the formation of neuromuscular synapses. Nonetheless, the size of synapses and the density of synaptic AChRs are reduced in AChR-␤ 3F/3F mutant mice. Moreover, synapses are structurally simplified and the organization of postjunctional folds is aberrant in mice lacking tyrosine phosphorylation of the AChR ␤-subunit. Furthermore, mutant AChRs cluster poorly in response to agrin and are readily extracted from the cell surface of cultured myotubes by non-ionic detergent. These data indicate that tyrosine phosphorylation of the AChR ␤-subunit has an important role in organizing AChRs and regulating synaptic differentiation.

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Tyrosine Phosphorylation of Nicotinic Acetylcholine Receptor Mediates Grb2 Binding

Cited by 45 publications

References 66 publications

Rapsyn carboxyl terminal domains mediate muscle specific kinase–induced phosphorylation of the muscle acetylcholine receptor

Rapsyn carboxyl terminal domains mediate muscle specific kinase–induced phosphorylation of the muscle acetylcholine receptor

Erbin Is a Protein Concentrated at Postsynaptic Membranes That Interacts with PSD-95

Synaptic differentiation is defective in mice lacking acetylcholine receptor β-subunit tyrosine phosphorylation

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