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PerleCan fix your muscle AChEs
Update
241
TRENDS in Neurosciences Vol.26 No.5 May 2003
PerleCan fix your muscle AChEs Michelle S. SteenStanley C. Froehner Department of Physiology and Biophysics, University of Washington, Seattle, WA 98195, USA
Efficient and accurate synaptic transmission requires proper localization of numerous signaling proteins in the synaptic membrane. At the neuromuscular junction, the nicotinic ACh receptor mediates postsynaptic depolarization, and acetylcholinesterase (AChE) terminates this process by hydrolyzing ACh. The mechanism by which the nerve directs receptor localization is understood in considerable detail; AChE clustering, by contrast, has received much less attention. Now, in a recent paper in Nature Neuroscience, the laboratories of Yoshiko Yamada and Richard Rotundo report that AChE clustering at the postsynaptic membrane requires perlecan, which binds both AChE and dystroglycan. A remarkably complex assembly, the synaptic form of acetylcholine esterase (AChE) contains 12 catalytic subunits (arranged as three tetramers). These are attached to a collagen-like tail (ColQ) [1] that links AChE to the synaptic basal lamina [2]. In humans, mutations in the colQ gene result in the absence of AChE from the neuromuscular synapse and cause congenital myasthenia gravis, a disorder of neuromuscular transmission [3]. Conversely, inhibiting AChE can treat the autoimmune form of myasthenia. In addition, neuromuscular synapse formation requires AChE. Zebrafish embryos lacking AChE enzymatic activity because of a point mutation have reduced ACh-receptor clustering [4]. Despite the importance of AChE in synaptic transmission, however, the exact molecular mechanism for AChE localization to the neuromuscular junction remains largely unknown. Clustering of AChE shares at least one mechanistic similarity with that of the ACh receptor: agrin clusters both [5]. At some point downstream of this initial step, however, the clustering mechanisms must diverge because ACh-receptor clustering does not require perlecan. Until recently, the protein interactions important for AChE anchoring were not known, but genetic analyses in mice and protein biochemistry have revealed two potentially important players: perlecan and a-dystroglycan, an important component of the dystrophin complex [6,7]. The dystrophin complex confers stability to the muscle membrane during muscle contractions. Mutations in several genes encoding dystrophin-complex proteins cause several different muscle wasting diseases, most notably Duchenne muscular dystrophy. Additionally, the dystrophin complex appears to play an important role in the development of the neuromuscular synapse. First, a-dystroglycan binds laminin and agrin – both proteins that induce ACh-receptor clustering [8,9]. Second, mouse Corresponding author: Stanley C. Froehner ([email protected]). http://tins.trends.com
mutants lacking specific members of the dystrophin complex (a-dystroglycan, a-syntrophin or a-dystrobrevin) display aberrantly developed neuromuscular junctions [7,10,11]. In some cases, the mutations perturb the levels and distributions of ACh receptors and AChE. However, the molecular means by which the dystrophin complex contributes to the proper formation of the postsynaptic membrane is unknown. Increasing evidence suggests that the complex serves as a scaffold for signaling molecules. In the absence of a-dystroglycan, perlecan and AChE do not colocalize with the ACh receptor, suggesting the requirement for the dystrophin complex in clustering AChE at the neuromuscular junction. Previous studies have also shown that a-dystroglycan binds to, and colocalizes with, perlecan, a heparan sulfate proteoglycan which, in turn, interacts and colocalizes with AChE [6] (Fig. 1). The recent findings of Arikawa-Hirasawa et al. provide strong evidence that the link between a-dystroglycan and AChE requires mediation by perlecan [12]. Perlecan deficiency in embryonic mice does not affect skeletal muscle development, nerve branching or fiber innervation, and it does not alter AChE expression. However, immunostaining for ColQ and the catalytic subunit of AChE revealed a complete absence of AChE from the neuromuscular junctions. Proper localization of AChE to the neuromuscular junction appears, therefore, to require perlecan. These interesting results raise new questions. a-Dystroglycan, like other members of the dystrophin complex, localizes not only to the postsynaptic membrane but also along the general sarcolemma. Thus, it is difficult to imagine how perlecan itself can direct AChE to the
ColQ
Perlecan α-Dystroglycan
Sarcoglycans
AChE Plasma membrane
Actin
β
α β γ δ Dystrobrevin Syn Syn
Dystrophin or utrophin Syn Syn
TRENDS in Neurosciences
Fig. 1. The collagen-like tail (ColQ) of acetylcholinesterase (AChE) binds carbohydrate chains of the heavily-glycosylated perlecan protein which, in turn, binds a-dystroglycan. Transmembrane b-dystroglycan associates with dystrophin or utrophin, thereby providing a link to the actin-based cytoskeleton. Other proteins of the dystrophin complex, including dystrobrevin, the syntrophins (Syn) and the sarcoglycans (a, b, g, and d), could play a signaling role important for muscle health.
Update
242
TRENDS in Neurosciences Vol.26 No.5 May 2003
synaptic membrane. In some cases, synaptic and nonsynaptic sites contain different isoforms of dystrophinassociated complex members. To date, however, we know of only a single gene that encodes dystroglycan. Several possibilities for the localization of perlecan and AChE at synapses come to mind. In view of the role of glycosylation in the interactions, perhaps differentially glycosylated forms of a-dystroglycan provide specificity to its interactions, synaptically and extra-synaptically. Because the collagen-tailed AChE is expressed locally and at high concentrations at the neuromuscular synapse, its ability to compete with other dystroglycan-binding proteins at the synapse might be a contributing factor. Selective intracellular assembly of the complex prior to externalization is another possibility. Finally other, yet to be identified, proteins might interact with AChE and contribute the specificity needed for synaptic localization. As we often find, interesting results lead to more challenging questions and the need for additional experimentation. References 1 Massoulie, J. et al. (1993) Molecular and cellular biology of cholinesterases. Prog. Neurobiol. 41, 31 – 91 2 Hall, Z.W. (1973) Multiple forms of acetylcholinesterase and their distribution in endplate and non-endplate regions of rat diaphragm muscle. J. Neurobiol. 4, 343 – 361
3 Ohno, K. et al. (2000) The spectrum of mutations causing end-plate acetylcholinesterase deficiency. Ann. Neurol. 47, 162 – 170 4 Behra, M. et al. (2002) Acetylcholinesterase is required for neuronal and muscular development in the zebrafish embryo. Nat. Neurosci. 5, 111 – 118 5 Wallace, B.G. (1989) Agrin-induced specializations contain cytoplasmic, membrane, and extracellular matrix-associated components of the postsynaptic apparatus. J. Neurosci. 9, 1294 – 1302 6 Peng, H.B. et al. (1999) Acetylcholinesterase clustering at the neuromuscular junction involves perlecan and dystroglycan. J. Cell Biol. 145, 911 – 921 7 Jacobson, C. et al. (2001) The dystroglycan complex is necessary for stabilization of acetylcholine receptor clusters at neuromuscular junctions and formation of the synaptic basement membrane. J. Cell Biol. 152, 435 – 450 8 Gee, S.H. et al. (1994) Dystroglycan-a, a dystrophin-associated glycoprotein, is a functional agrin receptor. Cell 77, 675– 686 9 Bowe, M.A. et al. (1994) Identification and purification of an agrin receptor from Torpedo postsynaptic membranes: a heteromeric complex related to the dystroglycans. Neuron 12, 1173 – 1180 10 Adams, M.E. et al. (2000) Absence of a-syntrophin leads to structurally aberrant neuromuscular synapses deficient in utrophin. J. Cell Biol. 150, 1385– 1398 11 Grady, R.M. et al. (2000) Maturation and maintenance of the neuromuscular synapse: genetic evidence for roles of the dystrophin – glycoprotein complex. Neuron 25, 279– 293 12 Arikawa-Hirasawa, E. et al. (2002) Absence of acetylcholinesterase at the neuromuscular junctions of perlecan-null mice. Nat. Neurosci. 5, 119 – 123 0166-2236/03/$ - see front matter q 2003 Elsevier Science Ltd. All rights reserved. doi:10.1016/S0166-2236(03)00077-8
Trends in Neurosciences: a forum for comment Controversial? Thought-provoking? If you wish to comment on articles published in Trends in Neurosciences, or would like to discuss issues of broad interest to neuroscientists, then why not write a Letter to the Editor? Letters should be up to 700 words. Please state clearly whether you wish the letter to be considered for publication. Letters are often sent to the author of the original article for their response, in which case both the letter and reply will be published together. Please note: submission does not guarantee publication
http://tins.trends.com
241
TRENDS in Neurosciences Vol.26 No.5 May 2003
PerleCan fix your muscle AChEs Michelle S. SteenStanley C. Froehner Department of Physiology and Biophysics, University of Washington, Seattle, WA 98195, USA
Efficient and accurate synaptic transmission requires proper localization of numerous signaling proteins in the synaptic membrane. At the neuromuscular junction, the nicotinic ACh receptor mediates postsynaptic depolarization, and acetylcholinesterase (AChE) terminates this process by hydrolyzing ACh. The mechanism by which the nerve directs receptor localization is understood in considerable detail; AChE clustering, by contrast, has received much less attention. Now, in a recent paper in Nature Neuroscience, the laboratories of Yoshiko Yamada and Richard Rotundo report that AChE clustering at the postsynaptic membrane requires perlecan, which binds both AChE and dystroglycan. A remarkably complex assembly, the synaptic form of acetylcholine esterase (AChE) contains 12 catalytic subunits (arranged as three tetramers). These are attached to a collagen-like tail (ColQ) [1] that links AChE to the synaptic basal lamina [2]. In humans, mutations in the colQ gene result in the absence of AChE from the neuromuscular synapse and cause congenital myasthenia gravis, a disorder of neuromuscular transmission [3]. Conversely, inhibiting AChE can treat the autoimmune form of myasthenia. In addition, neuromuscular synapse formation requires AChE. Zebrafish embryos lacking AChE enzymatic activity because of a point mutation have reduced ACh-receptor clustering [4]. Despite the importance of AChE in synaptic transmission, however, the exact molecular mechanism for AChE localization to the neuromuscular junction remains largely unknown. Clustering of AChE shares at least one mechanistic similarity with that of the ACh receptor: agrin clusters both [5]. At some point downstream of this initial step, however, the clustering mechanisms must diverge because ACh-receptor clustering does not require perlecan. Until recently, the protein interactions important for AChE anchoring were not known, but genetic analyses in mice and protein biochemistry have revealed two potentially important players: perlecan and a-dystroglycan, an important component of the dystrophin complex [6,7]. The dystrophin complex confers stability to the muscle membrane during muscle contractions. Mutations in several genes encoding dystrophin-complex proteins cause several different muscle wasting diseases, most notably Duchenne muscular dystrophy. Additionally, the dystrophin complex appears to play an important role in the development of the neuromuscular synapse. First, a-dystroglycan binds laminin and agrin – both proteins that induce ACh-receptor clustering [8,9]. Second, mouse Corresponding author: Stanley C. Froehner ([email protected]). http://tins.trends.com
mutants lacking specific members of the dystrophin complex (a-dystroglycan, a-syntrophin or a-dystrobrevin) display aberrantly developed neuromuscular junctions [7,10,11]. In some cases, the mutations perturb the levels and distributions of ACh receptors and AChE. However, the molecular means by which the dystrophin complex contributes to the proper formation of the postsynaptic membrane is unknown. Increasing evidence suggests that the complex serves as a scaffold for signaling molecules. In the absence of a-dystroglycan, perlecan and AChE do not colocalize with the ACh receptor, suggesting the requirement for the dystrophin complex in clustering AChE at the neuromuscular junction. Previous studies have also shown that a-dystroglycan binds to, and colocalizes with, perlecan, a heparan sulfate proteoglycan which, in turn, interacts and colocalizes with AChE [6] (Fig. 1). The recent findings of Arikawa-Hirasawa et al. provide strong evidence that the link between a-dystroglycan and AChE requires mediation by perlecan [12]. Perlecan deficiency in embryonic mice does not affect skeletal muscle development, nerve branching or fiber innervation, and it does not alter AChE expression. However, immunostaining for ColQ and the catalytic subunit of AChE revealed a complete absence of AChE from the neuromuscular junctions. Proper localization of AChE to the neuromuscular junction appears, therefore, to require perlecan. These interesting results raise new questions. a-Dystroglycan, like other members of the dystrophin complex, localizes not only to the postsynaptic membrane but also along the general sarcolemma. Thus, it is difficult to imagine how perlecan itself can direct AChE to the
ColQ
Perlecan α-Dystroglycan
Sarcoglycans
AChE Plasma membrane
Actin
β
α β γ δ Dystrobrevin Syn Syn
Dystrophin or utrophin Syn Syn
TRENDS in Neurosciences
Fig. 1. The collagen-like tail (ColQ) of acetylcholinesterase (AChE) binds carbohydrate chains of the heavily-glycosylated perlecan protein which, in turn, binds a-dystroglycan. Transmembrane b-dystroglycan associates with dystrophin or utrophin, thereby providing a link to the actin-based cytoskeleton. Other proteins of the dystrophin complex, including dystrobrevin, the syntrophins (Syn) and the sarcoglycans (a, b, g, and d), could play a signaling role important for muscle health.
Update
242
TRENDS in Neurosciences Vol.26 No.5 May 2003
synaptic membrane. In some cases, synaptic and nonsynaptic sites contain different isoforms of dystrophinassociated complex members. To date, however, we know of only a single gene that encodes dystroglycan. Several possibilities for the localization of perlecan and AChE at synapses come to mind. In view of the role of glycosylation in the interactions, perhaps differentially glycosylated forms of a-dystroglycan provide specificity to its interactions, synaptically and extra-synaptically. Because the collagen-tailed AChE is expressed locally and at high concentrations at the neuromuscular synapse, its ability to compete with other dystroglycan-binding proteins at the synapse might be a contributing factor. Selective intracellular assembly of the complex prior to externalization is another possibility. Finally other, yet to be identified, proteins might interact with AChE and contribute the specificity needed for synaptic localization. As we often find, interesting results lead to more challenging questions and the need for additional experimentation. References 1 Massoulie, J. et al. (1993) Molecular and cellular biology of cholinesterases. Prog. Neurobiol. 41, 31 – 91 2 Hall, Z.W. (1973) Multiple forms of acetylcholinesterase and their distribution in endplate and non-endplate regions of rat diaphragm muscle. J. Neurobiol. 4, 343 – 361
3 Ohno, K. et al. (2000) The spectrum of mutations causing end-plate acetylcholinesterase deficiency. Ann. Neurol. 47, 162 – 170 4 Behra, M. et al. (2002) Acetylcholinesterase is required for neuronal and muscular development in the zebrafish embryo. Nat. Neurosci. 5, 111 – 118 5 Wallace, B.G. (1989) Agrin-induced specializations contain cytoplasmic, membrane, and extracellular matrix-associated components of the postsynaptic apparatus. J. Neurosci. 9, 1294 – 1302 6 Peng, H.B. et al. (1999) Acetylcholinesterase clustering at the neuromuscular junction involves perlecan and dystroglycan. J. Cell Biol. 145, 911 – 921 7 Jacobson, C. et al. (2001) The dystroglycan complex is necessary for stabilization of acetylcholine receptor clusters at neuromuscular junctions and formation of the synaptic basement membrane. J. Cell Biol. 152, 435 – 450 8 Gee, S.H. et al. (1994) Dystroglycan-a, a dystrophin-associated glycoprotein, is a functional agrin receptor. Cell 77, 675– 686 9 Bowe, M.A. et al. (1994) Identification and purification of an agrin receptor from Torpedo postsynaptic membranes: a heteromeric complex related to the dystroglycans. Neuron 12, 1173 – 1180 10 Adams, M.E. et al. (2000) Absence of a-syntrophin leads to structurally aberrant neuromuscular synapses deficient in utrophin. J. Cell Biol. 150, 1385– 1398 11 Grady, R.M. et al. (2000) Maturation and maintenance of the neuromuscular synapse: genetic evidence for roles of the dystrophin – glycoprotein complex. Neuron 25, 279– 293 12 Arikawa-Hirasawa, E. et al. (2002) Absence of acetylcholinesterase at the neuromuscular junctions of perlecan-null mice. Nat. Neurosci. 5, 119 – 123 0166-2236/03/$ - see front matter q 2003 Elsevier Science Ltd. All rights reserved. doi:10.1016/S0166-2236(03)00077-8
Trends in Neurosciences: a forum for comment Controversial? Thought-provoking? If you wish to comment on articles published in Trends in Neurosciences, or would like to discuss issues of broad interest to neuroscientists, then why not write a Letter to the Editor? Letters should be up to 700 words. Please state clearly whether you wish the letter to be considered for publication. Letters are often sent to the author of the original article for their response, in which case both the letter and reply will be published together. Please note: submission does not guarantee publication
http://tins.trends.com