The Experts below are selected from a list of 1461 Experts worldwide ranked by ideXlab platform
Jane E. Hewitt - One of the best experts on this subject based on the ideXlab platform.
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A new monoclonal antibody against human Alpha-Dystroglycan has potential diagnostic applications
Neuromuscular Disorders, 2014Co-Authors: Emma L. Humphrey, Francesca Sciandra, Andrea Brancaccio, E. Lacey, L.t. Le, Christopher Morris, Jane E. Hewitt, Ian Holt, Rita Barresi, Caroline SewryAbstract:We have generated novel monoclonal antibodies (mAbs) against Alpha-Dystroglycan to immunolabel the sarcolemma in human muscle biopsies. For one of these, DAG-6F4, a seven amino-acid epitope, PNQRPEL, was identified using phage-displayed peptides and is located immediately after the highly-glycosylated mucin domain of Alpha-Dystroglycan. On western blots of recombinant Alpha-Dystroglycan, epitope accessibility was reduced, but not entirely prevented, by glycosylation. DAG-6F4 immunolabelling was markedly reduced in muscle biopsies from Duchenne muscular dystrophy patients consistent with a disruption in the Dystroglycan complex. In a range of Dystroglycanopathy patients with reduced/altered glycosylation, staining by DAG-6F4 was generally less reduced than staining by IIH6 (an antibody commonly used to identify glycosylated Alpha-Dystroglycan), though the extent of the differences between the two antibodies varied between different patients, some biopsies showing reductions in core protein as well as its glycosylation. A second mAb, DAG-3H9, gave similar results, but recognised a different epitope, GDRAP, closer to the C-terminus. There are currently few antibodies available against core Alpha-Dystroglycan, so DAG-6F4 represents a useful addition to the antibody repertoire for evaluating the Dystroglycan complex in neuromuscular disorders.
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A new monoclonal antibody DAG-6F4 against human Alpha-Dystroglycan reveals reduced core protein in some, but not all, Dystroglycanopathy patients.
Neuromuscular disorders : NMD, 2014Co-Authors: Emma L. Humphrey, Lucy Feng, Francesca Sciandra, Andrea Brancaccio, E. Lacey, Jane E. Hewitt, Ian Holt, Charlotte R. Morris, Rita BarresiAbstract:We generated a novel monoclonal antibody, DAG-6F4, against Alpha-Dystroglycan which immunolabels the sarcolemma in human muscle biopsies. Its seven amino-acid epitope, PNQRPEL, was identified using phage-displayed peptides and is located immediately after the highly-glycosylated mucin domain of Alpha-Dystroglycan. On Western blots of recombinant Alpha-Dystroglycan, epitope accessibility was reduced, but not entirely prevented, by glycosylation. DAG-6F4 immunolabelling was markedly reduced in muscle biopsies from Duchenne muscular dystrophy patients consistent with disruption of the Dystroglycan complex. In a range of Dystroglycanopathy patients with reduced/altered glycosylation, staining by DAG-6F4 was often less reduced than staining by IIH6 (antibody against the glycan epitope added by LARGE and commonly used to identify glycosylated Alpha-Dystroglycan). Whereas IIH6 was reduced in all patients, DAG-6F4 was hardly changed in a LARGE patient, less reduced than IIH6 in limb-girdle muscular dystrophy type 2I, but as reduced as IIH6 in some congenital muscular dystrophy patients. Although absence of the LARGE-dependent laminin-binding site appears not to affect Alpha-Dystroglycan stability at the sarcolemma, the results suggest that further reduction in aDG glycosylation may reduce its stability. These studies suggest that DAG-6F4 may be a useful addition to the antibody repertoire for evaluating the Dystroglycan complex in neuromuscular disorders.
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P.1.17 Immunostaining of the sarcolemma with a new monoclonal antibody against Alpha-Dystroglycan core and its relevance to diagnosis
Neuromuscular Disorders, 2013Co-Authors: E. Wilson, Kazuhiro Kobayashi, Tatsushi Toda, Francesca Sciandra, Andrea Brancaccio, E. Lacey, Jane E. Hewitt, Ian Holt, Caroline Sewry, Susan C. BrownAbstract:Alpha-Dystroglycan ( α DG) is an extracellular, glycosylated protein attached to the outer plasma membrane by its interaction with the transmembrane protein, beta-Dystroglycan. Genetic mutations that alter the glycosylation state of α DG cause a number of inherited disorders, including Fukuyama MD and certain congenital and limb-girdle MDs. Many of the pathogenic mutations are in proteins that carry out glycosylation of α DG and not in the DG gene itself. These enzymes are usually located in the Golgi where proteins are modified before export to the extracellular space. Amino-acid mutations in α DG itself can also cause congenital MD when sugar attachment to the α DG backbone is affected. Antibodies against Dystroglycans immunostain the sarcolemma in a similar manner to dystrophin antibodies. It would be useful to the muscle pathologist to have monoclonal antibodies (mAbs) that recognize specific glycosylated forms of α DG associated with different mutations or disease states, since such mAbs would be useful for diagnosis using muscle biopsy sections. There are problems of reproducibility with existing mAbs against α DG, but it is still unclear whether those problems are attributable to the antibodies themselves or to the nature of the glycosylated epitopes on α DG. To determine whether it is possible to obtain improved mAbs for diagnostic use, we have begun a collaborative programme, supported by LGMD2I and CureCMD, to produce new mAbs against glycosylated and non-glycosylated forms of α DG. As a first step, we now describe a new mAb raised against bacterial recombinant α DG core sequence, which gives good staining of the sarcolemma in human muscle biopsy sections. We have mapped the epitope recognized by this mAb to short sequence that appears to be accessible to antibody in BOTH glycosylated and non-glycosylated forms of α DG.
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Post-translational modification of {Alpha}-Dystroglycan is not critical for lymphocytic choriomeningitis virus receptor function in vivo; ; The journal of general virology : JGV;
2008Co-Authors: Mauro Imperiali, Jane E. Hewitt, Roman Spörri, Annette OxeniusAbstract:{Alpha}-Dystroglycan ({Alpha}-DG) is a ubiquitously expressed molecule that has been identified as a cellular receptor for lymphocytic choriomeningitis virus (LCMV) and other arenaviruses. Recently, it was demonstrated that LCMV receptor function is critically dependent on post-translational modifications, namely glycosylation. In particular, it was shown that O-mannosylation, a rare type of mammalian O-linked glycosylation, is important in determining the binding of LCMV to its cellular receptor. All studies carried out so far showed a dependence on glycosylation in LCMV receptor function in vitro. This work extended these studies to two in vivo models of {Alpha}-DG hypoglycosylation. The results confirm earlier findings on the in vitro dependence of carbohydrate modifications in LCMV receptor function. However, experiments in animal models showed that this dependence was only very weak in vivo. It is likely that alternative receptors or alternative entry pathways may account for this attenuated in vivo phenotype.
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post translational modification of Alpha Dystroglycan is not critical for lymphocytic choriomeningitis virus receptor function in vivo the journal of general virology jgv
The journal of general virology : JGV, 2008Co-Authors: Mauro Imperiali, Jane E. Hewitt, Roman Spa Rri, Annette OxeniusAbstract:{Alpha}-Dystroglycan ({Alpha}-DG) is a ubiquitously expressed molecule that has been identified as a cellular receptor for lymphocytic choriomeningitis virus (LCMV) and other arenaviruses. Recently, it was demonstrated that LCMV receptor function is critically dependent on post-translational modifications, namely glycosylation. In particular, it was shown that O-mannosylation, a rare type of mammalian O-linked glycosylation, is important in determining the binding of LCMV to its cellular receptor. All studies carried out so far showed a dependence on glycosylation in LCMV receptor function in vitro. This work extended these studies to two in vivo models of {Alpha}-DG hypoglycosylation. The results confirm earlier findings on the in vitro dependence of carbohydrate modifications in LCMV receptor function. However, experiments in animal models showed that this dependence was only very weak in vivo. It is likely that alternative receptors or alternative entry pathways may account for this attenuated in vivo phenotype.
Andrea Brancaccio - One of the best experts on this subject based on the ideXlab platform.
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Analysis of α-Dystroglycan/LG Domain Binding Modes: Investigating Protein Motifs That Regulate the Affinity of Isolated LG Domains.
Frontiers in molecular biosciences, 2019Co-Authors: Christopher E. Dempsey, Maria Giulia Bigotti, Josephine C. Adams, Andrea BrancaccioAbstract:Dystroglycan (DG) is an adhesion complex that links the cytoskeleton to the surrounding extracellular matrix in skeletal muscle and a wide variety of other tissues. It is composed of a highly glycosylated extracellular Alpha-DG associated noncovalently with a transmembrane beta-DG whose cytodomain interacts with dystrophin and its isoforms. Alpha-Dystroglycan (Alpha-DG) binds tightly and in a calcium-dependent fashion to multiple extracellular proteins and proteoglycans, each of which harbours at least one, or, more frequently, tandem arrays of laminin-globular (LG) domains. Considerable biochemical and structural work has accumulated on the Alpha-DG-binding LG domains, highlighting a significant heterogeneity in ligand-binding properties of domains from different proteins as well as between single and multiple LG domains within the same protein. Here we review biochemical, structural and functional information on the LG domains reported to bind Alpha-Dystroglycan. In addition, we have incorporated bioinformatics and modelling to explore whether specific motifs responsible for Alpha-Dystroglycan recognition can be identified within isolated LG domains. In particular, we analysed the LG domains of slits and agrin as well as those of paradigmatic Alpha-DG non-binders such as laminin-Alpha3. While some stretches of basic residues may be important, no universally conserved motifs could be identified. However, the data confirm that the coordinated calcium atom within the LG domain is needed to establish an interaction with the sugars of Alpha-DG, although it appears that this alone is insufficient to mediate significant Alpha-DG binding. We develop a scenario involving different binding modes of a single LG domain unit, or tandemly repeated units, with Alpha-DG. A variability of binding modes might be important to generate a range of affinities to allow physiological regulation of this interaction, reflecting its crucial biological importance.
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A new monoclonal antibody against human Alpha-Dystroglycan has potential diagnostic applications
Neuromuscular Disorders, 2014Co-Authors: Emma L. Humphrey, Francesca Sciandra, Andrea Brancaccio, E. Lacey, L.t. Le, Christopher Morris, Jane E. Hewitt, Ian Holt, Rita Barresi, Caroline SewryAbstract:We have generated novel monoclonal antibodies (mAbs) against Alpha-Dystroglycan to immunolabel the sarcolemma in human muscle biopsies. For one of these, DAG-6F4, a seven amino-acid epitope, PNQRPEL, was identified using phage-displayed peptides and is located immediately after the highly-glycosylated mucin domain of Alpha-Dystroglycan. On western blots of recombinant Alpha-Dystroglycan, epitope accessibility was reduced, but not entirely prevented, by glycosylation. DAG-6F4 immunolabelling was markedly reduced in muscle biopsies from Duchenne muscular dystrophy patients consistent with a disruption in the Dystroglycan complex. In a range of Dystroglycanopathy patients with reduced/altered glycosylation, staining by DAG-6F4 was generally less reduced than staining by IIH6 (an antibody commonly used to identify glycosylated Alpha-Dystroglycan), though the extent of the differences between the two antibodies varied between different patients, some biopsies showing reductions in core protein as well as its glycosylation. A second mAb, DAG-3H9, gave similar results, but recognised a different epitope, GDRAP, closer to the C-terminus. There are currently few antibodies available against core Alpha-Dystroglycan, so DAG-6F4 represents a useful addition to the antibody repertoire for evaluating the Dystroglycan complex in neuromuscular disorders.
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A new monoclonal antibody DAG-6F4 against human Alpha-Dystroglycan reveals reduced core protein in some, but not all, Dystroglycanopathy patients.
Neuromuscular disorders : NMD, 2014Co-Authors: Emma L. Humphrey, Lucy Feng, Francesca Sciandra, Andrea Brancaccio, E. Lacey, Jane E. Hewitt, Ian Holt, Charlotte R. Morris, Rita BarresiAbstract:We generated a novel monoclonal antibody, DAG-6F4, against Alpha-Dystroglycan which immunolabels the sarcolemma in human muscle biopsies. Its seven amino-acid epitope, PNQRPEL, was identified using phage-displayed peptides and is located immediately after the highly-glycosylated mucin domain of Alpha-Dystroglycan. On Western blots of recombinant Alpha-Dystroglycan, epitope accessibility was reduced, but not entirely prevented, by glycosylation. DAG-6F4 immunolabelling was markedly reduced in muscle biopsies from Duchenne muscular dystrophy patients consistent with disruption of the Dystroglycan complex. In a range of Dystroglycanopathy patients with reduced/altered glycosylation, staining by DAG-6F4 was often less reduced than staining by IIH6 (antibody against the glycan epitope added by LARGE and commonly used to identify glycosylated Alpha-Dystroglycan). Whereas IIH6 was reduced in all patients, DAG-6F4 was hardly changed in a LARGE patient, less reduced than IIH6 in limb-girdle muscular dystrophy type 2I, but as reduced as IIH6 in some congenital muscular dystrophy patients. Although absence of the LARGE-dependent laminin-binding site appears not to affect Alpha-Dystroglycan stability at the sarcolemma, the results suggest that further reduction in aDG glycosylation may reduce its stability. These studies suggest that DAG-6F4 may be a useful addition to the antibody repertoire for evaluating the Dystroglycan complex in neuromuscular disorders.
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Insights from Molecular Dynamics Simulations: Structural Basis for the V567D Mutation-Induced Instability of Zebrafish Alpha-Dystroglycan and Comparison with the Murine Model
2014Co-Authors: Davide Pirolli, Francesca Sciandra, Bruno Giardina, Andrea Brancaccio, Manuela Bozzi, Maria Cristina De RosaAbstract:A missense amino acid mutation of valine to aspartic acid in 567 position of Alpha-Dystroglycan (DG), identified in dag1-mutated zebrafish, results in a reduced transcription and a complete absence of the protein. Lacking experimental structural data for zebrafish DG domains, the detailed mechanism for the observed mutation-induced destabilization of the DG complex and membrane damage, remained unclear. With the aim to contribute to a better clarification of the structure-function relationships featuring the DG complex, three-dimensional structural models of wild-type and mutant (V567D) C-terminal domain of Alpha-DG from zebrafish were constructed by a template-based modelling approach. We then ran extensive molecular dynamics (MD) simulations to reveal the structural and dynamic properties of the C-terminal domain and to evaluate the effect of the single mutation on Alpha-DG stability. A comparative study has been also carried out on our previously generated model of murine Alpha-DG C-terminal domain including the I591D mutation, which is topologically equivalent to the V567D mutation found in zebrafish. Trajectories from MD simulations were analyzed in detail, revealing extensive structural disorder involving multiple beta-strands in the mutated variant of the zebrafish protein whereas local effects have been detected in the murine protein. A biochemical analysis of the murine Alpha-DG mutant I591D confirmed a pronounced instability of the protein. Taken together, the computational and biochemical analysis suggest that the V567D/I591D mutation, belonging to the G beta-strand, plays a key role in inducing a destabilization of the Alpha-DG C-terminal Ig-like domain that could possibly affect and propagate to the entire DG complex. The structural features herein identified may be of crucial help to understand the molecular basis of primary Dystroglycanopathies.
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P.1.17 Immunostaining of the sarcolemma with a new monoclonal antibody against Alpha-Dystroglycan core and its relevance to diagnosis
Neuromuscular Disorders, 2013Co-Authors: E. Wilson, Kazuhiro Kobayashi, Tatsushi Toda, Francesca Sciandra, Andrea Brancaccio, E. Lacey, Jane E. Hewitt, Ian Holt, Caroline Sewry, Susan C. BrownAbstract:Alpha-Dystroglycan ( α DG) is an extracellular, glycosylated protein attached to the outer plasma membrane by its interaction with the transmembrane protein, beta-Dystroglycan. Genetic mutations that alter the glycosylation state of α DG cause a number of inherited disorders, including Fukuyama MD and certain congenital and limb-girdle MDs. Many of the pathogenic mutations are in proteins that carry out glycosylation of α DG and not in the DG gene itself. These enzymes are usually located in the Golgi where proteins are modified before export to the extracellular space. Amino-acid mutations in α DG itself can also cause congenital MD when sugar attachment to the α DG backbone is affected. Antibodies against Dystroglycans immunostain the sarcolemma in a similar manner to dystrophin antibodies. It would be useful to the muscle pathologist to have monoclonal antibodies (mAbs) that recognize specific glycosylated forms of α DG associated with different mutations or disease states, since such mAbs would be useful for diagnosis using muscle biopsy sections. There are problems of reproducibility with existing mAbs against α DG, but it is still unclear whether those problems are attributable to the antibodies themselves or to the nature of the glycosylated epitopes on α DG. To determine whether it is possible to obtain improved mAbs for diagnostic use, we have begun a collaborative programme, supported by LGMD2I and CureCMD, to produce new mAbs against glycosylated and non-glycosylated forms of α DG. As a first step, we now describe a new mAb raised against bacterial recombinant α DG core sequence, which gives good staining of the sarcolemma in human muscle biopsy sections. We have mapped the epitope recognized by this mAb to short sequence that appears to be accessible to antibody in BOTH glycosylated and non-glycosylated forms of α DG.
Markus A. Rüegg - One of the best experts on this subject based on the ideXlab platform.
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G.P.212
Neuromuscular Disorders, 2014Co-Authors: Sarina Meinen, Geraldine Maier, M. Chauhan, Shuo Lin, Karen K. Mckee, Stephanie C. Crosson, Peter D. Yurchenco, Markus A. RüeggAbstract:The basement membrane surrounding skeletal muscle fibers (myomatrix) and its binding to the sarcolemma is important for the structural stability of muscle. The myomatrix consists of laminin-211 (Lm-211), collagen IV and several associated components including nidogen and perlecan. Assembly of the myomatrix is initiated by Lm-211 that binds to its cell surface receptors (Alpha-Dystroglycan and integrins) and then creates a primary scaffold by self-assembly. Mutations in the α2 chain of Lm-211 result in a severe form of congenital muscular dystrophy, called MDC1A. In MDC1A patients and the dyW/dyW mouse model expression of Lm-411 is increased as an attempt to compensate for the loss of Lm-211. However, Lm-411 cannot self-assemble and binds only weakly to the sarcolemma. Previous proof-of-principle studies in dyW/dyW mice have shown that transgenic expression of mini-agrin, which binds to Lm-411 and to Alpha-Dystroglycan, strongly ameliorates the dystrophic phenotype. We here present data that introduction of a transgene that allows self-polymerization of Lm-411 has some, although rather weak therapeutic activity in dyW/dyW mice. However, when combined with mini-agrin, this construct shows a strong synergistic effect. The improvements are seen on all levels, such as behavior, weight gain and the histology of skeletal muscles. We are currently evaluating its effect on muscle force and survival. Moreover, we analyze the signaling pathways involved in the amelioration. We also plan to test a new construct that combines the laminin polymerization and the Alpha-Dystroglycan binding regions. In summary, our data provide formal proof that MDC1A is caused by defects in myomatrix assembly and its connection to the sarcolemma. Our data will have significant impact in future attempts to develop new treatment strategies for MDC1A.
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Activation of muscle-specific receptor tyrosine kinase and binding to Dystroglycan are regulated by alternative mRNA splicing of agrin
The Journal of biological chemistry, 2006Co-Authors: Patrick Scotton, Francesca Sciandra, Andrea Brancaccio, Thomas Meier, Dorothee Bleckmann, Michael Stebler, Jörg Stetefeld, Markus A. RüeggAbstract:Agrin induces the aggregation of postsynaptic proteins at the neuromuscular junction (NMJ). This activity requires the receptor-tyrosine kinase MuSK. Agrin isoforms differ in short amino acid stretches at two sites, called A and B, that are localized in the two most C-terminal laminin G (LG) domains. Importantly, agrin isoforms greatly differ in their activities of inducing MuSK phosphorylation and of binding to Alpha-Dystroglycan. By using site-directed mutagenesis, we characterized the amino acids important for these activities of agrin. We find that the conserved tripeptide asparagineglutamate-isoleucine in the eight-amino acid long insert at the B-site is necessary and sufficient for full MuSK phosphorylation activity. However, even if all eight amino acids were replaced by alanines, this agrin mutant still has significantly higher MuSK phosphorylation activity than the splice version lacking any insert. We also show that binding to Alpha-Dystroglycan requires at least two LG domains and that amino acid inserts at the A and the B splice sites negatively affect binding.
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Dystroglycan is a dual receptor for agrin and laminin 2 in schwann cell membrane
Journal of Biological Chemistry, 1996Co-Authors: Hiroki Yamada, Teruo Shimizu, Louise V. B. Anderson, Hisae Hori, Markus A. Rüegg, Alain J Denzer, Takeshi Tanaka, Sachiko Fujita, Hiroko Fukutaohi, Kiichiro MatsumuraAbstract:We have shown previously that Alpha-Dystroglycan with a molecular mass of 120 kDa is a Schwann cell receptor of laminin-2, the endoneurial isoform of laminin comprised of the Alpha2, beta1, and gamma1 chains. In this paper, we show that Schwann cell Alpha-Dystroglycan is also a receptor of agrin, an acetylcholine receptor-aggregating molecule having partial homology to laminin Alpha chains in the C terminus. Immunochemical analysis demonstrates that the peripheral nerve isoform of agrin is a 400-kDa component of the endoneurial basal lamina and is co-localized with Alpha-Dystroglycan surrounding the outermost layer of myelin sheath of peripheral nerve fibers. Blot overlay analysis demonstrates that both endogenous peripheral nerve agrin and laminin-2 bind to Schwann cell Alpha-Dystroglycan. Recombinant C-terminal fragment of the peripheral nerve isoform of agrin also binds to Schwann cell Alpha-Dystroglycan, confirming that the binding site for Schwann cell Alpha-Dystroglycan resides in the C terminus of agrin molecule. Furthermore, the binding of recombinant agrin C-terminal fragment to Schwann cell Alpha-Dystroglycan competes with that of laminin-2. All together, these results indicate that Alpha-Dystroglycan is a dual receptor for agrin and laminin-2 in the Schwann cell membrane.
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AChR phosphorylation and aggregation induced by an agrin fragment that lacks the binding domain for Alpha-Dystroglycan.
The EMBO journal, 1996Co-Authors: Thomas Meier, Matthias Gesemann, Valeria Cavalli, Markus A. Rüegg, Bruce G. WallaceAbstract:Agrin induces both phosphorylation and aggregation of nicotinic acetylcholine receptors (AChRs) when added to myotubes in culture, apparently by binding to a specific receptor on the myotube surface. One such agrin receptor is Alpha-Dystroglycan, although binding to Alpha-Dystroglycan appears not to mediate AChR aggregation. To determine whether agrin-induced AChR phosphorylation is mediated by Alpha-Dystroglycan or by a different agrin receptor, fragments of recombinant agrin that differ in affinity for Alpha-Dystroglycan were examined for their ability to induce AChR phosphorylation and aggregation in mouse C2 myotubes. The carboxy-terminal 95 kDa agrin fragment agrin-c95(A0B0), which binds to Alpha-Dystroglycan with high affinity, failed to induce AChR phosphorylation and aggregation. In contrast, agrin-c95(A4B8) which binds less strongly to Alpha-Dystroglycan, induced both phosphorylation and aggregation, as did a small 21 kDa fragment of agrin, agrin-c21(B8), that completely lacks the binding domain for Alpha-Dystroglycan. We conclude that agrin-induced AChR phosphorylation and aggregation are triggered by an agrin receptor that is distinct from Alpha-Dystroglycan.
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Alternative Splicing of Agrin Alters Its Binding to Heparin, Dystroglycan, and the Putative Agrin Receptor
Neuron, 1996Co-Authors: Matthias Gesemann, Andrea Brancaccio, Valeria Cavalli, Alain J Denzer, Beat Schumacher, Markus A. RüeggAbstract:Agrin is a heparan sulfate proteoglycan that induces aggregation of acetylcholine receptors (AChRs) at the neuromuscular synapse. This aggregating activity is modulated by alternative splicing. Here, we compared binding of agrin isoforms to heparin, Alpha-Dystroglycan, and cultured myotubes. We find that the alternatively spliced 4 amino acids insert (KSRK) is required for heparin binding. The binding affinity of agrin isoforms to Alpha-Dystroglycan correlates neither with binding to heparin nor with their AChR-aggregating activities. Moreover, the minimal fragment sufficient to induce AChR aggregation does not bind to Alpha-Dystroglycan. Nevertheless, this fragment still binds to cultured muscle cells. Its binding is completed only by agrin isoforms that are active in AChR aggregation, and therefore this binding site is likely to represent the receptor that initiates AChR clustering.
Steven A. Moore - One of the best experts on this subject based on the ideXlab platform.
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cardiac pathology exceeds skeletal muscle pathology in two cases of limb girdle muscular dystrophy type 2i
Muscle & Nerve, 2009Co-Authors: Marta Margeta, Anne M. Connolly, Thomas L. Winder, Alan Pestronk, Steven A. MooreAbstract:Limb-girdle muscular dystrophy type 2I (LGMD-2I) is caused by mutations in the fukutin-related protein gene (FKRP) that lead to abnormal glycosylation of Alpha-Dystroglycan in skeletal muscle. Heart involvement in LGMD-2I is common, but little is known about a underlying cardiac pathology. Herein we describe two patients with LGMD-2I (homozygous FKRP mutation c.826C>A, p.Leu276Ile) who developed severe congestive heart failure that required cardiac transplantation. The dystrophic pathology and impairment of Alpha-Dystroglycan glycosylation were severe in the heart but mild in skeletal muscle, underscoring the lack of correlation between cardiac and skeletal muscle involvement in some LGMD-2I patients.
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basal lamina strengthens cell membrane integrity via the laminin g domain binding motif of α Dystroglycan
Proceedings of the National Academy of Sciences of the United States of America, 2009Co-Authors: Renzhi Han, Steven A. Moore, Takako Yoshidamoriguchi, Stefan Kunz, Motoi Kanagawa, Daniel E. Michele, Erik P Rader, David E Muirhead, Susan T Iannaccone, Katsuya MiyakeAbstract:Skeletal muscle basal lamina is linked to the sarcolemma through transmembrane receptors, including integrins and Dystroglycan. The function of Dystroglycan relies critically on posttranslational glycosylation, a common target shared by a genetically heterogeneous group of muscular dystrophies characterized by Alpha-Dystroglycan hypoglycosylation. Here we show that both Dystroglycan and integrin Alpha7 contribute to force-production of muscles, but that only disruption of Dystroglycan causes detachment of the basal lamina from the sarcolemma and renders muscle prone to contraction-induced injury. These phenotypes of Dystroglycan-null muscles are recapitulated by Large(myd) muscles, which have an intact dystrophin-glycoprotein complex and lack only the laminin globular domain-binding motif on Alpha-Dystroglycan. Compromised sarcolemmal integrity is directly shown in Large(myd) muscles and similarly in normal muscles when arenaviruses compete with matrix proteins for binding Alpha-Dystroglycan. These data provide direct mechanistic insight into how the Dystroglycan-linked basal lamina contributes to the maintenance of sarcolemmal integrity and protects muscles from damage.
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Cardiac pathology exceeds skeletal muscle pathology in two cases of limb-girdle muscular dystrophy type 2I†
Muscle & Nerve, 2009Co-Authors: Marta Margeta, Anne M. Connolly, Thomas L. Winder, Alan Pestronk, Steven A. MooreAbstract:Limb-girdle muscular dystrophy type 2I (LGMD-2I) is caused by mutations in the fukutin-related protein gene (FKRP) that lead to abnormal glycosylation of Alpha-Dystroglycan in skeletal muscle. Heart involvement in LGMD-2I is common, but little is known about a underlying cardiac pathology. Herein we describe two patients with LGMD-2I (homozygous FKRP mutation c.826C>A, p.Leu276Ile) who developed severe congestive heart failure that required cardiac transplantation. The dystrophic pathology and impairment of Alpha-Dystroglycan glycosylation were severe in the heart but mild in skeletal muscle, underscoring the lack of correlation between cardiac and skeletal muscle involvement in some LGMD-2I patients.
Kevin P. Campbell - One of the best experts on this subject based on the ideXlab platform.
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220th ENMC workshop: Dystroglycan and the Dystroglycanopathies Naarden, The Netherlands, 27-29 May 2016.
Neuromuscular disorders : NMD, 2016Co-Authors: Susan C. Brown, Kevin P. Campbell, Steve J. Winder, Neil Bevan, Lyndsey Bevan, Sue Brown, Hans Van Bokhoven, Sebahattin Cirak, Bulmaro Cisneros, Holly ColognatoAbstract:Highlights • Review of clinical phenotypes associated with the Dystroglycanopathies. • Discussion of current animal models and their contribution to understanding the disease process. • New insight into the glycosylation of Alpha Dystroglycan and the role of LARGE. • Structural information on the LARGE glycan.
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Decoding Arenavirus Pathogenesis: Essential Roles for Alpha-Dystroglycan-Virus Interactions and the Immune Response
Virology, 2010Co-Authors: Kevin P. CampbellAbstract:Pathogenesis following a virus infection results from interactions between the virus and its host. The outcome is determined by tipping the balance between virulence of the virus or susceptibility/resistance of the host to favor one or the other. This review focuses on two important members of the Old World arenavirus family: Lassa fever virus (LFV), a robust human pathogen that causes a severe acute hemorrhagic disease; and lymphocytic choriomeningitis virus (LCMV), also a human pathogen but better known in the context of its rodent model. Research with this model has uncovered and illuminated many of our current concepts in immunobiology and viral pathogenesis. Presented here are recent advances that form the framework for a better understanding of how viruses induce and maintain persistent infection as well as for the pathogenesis associated with acute LFV infection. A major component for understanding the pathogenesis of these arenaviruses revolves around study of the interaction of virus with its receptor, Alpha-Dystroglycan (α-DG).
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Abstract 996: Loss of Alpha-Dystroglycan laminin binding in epithelium-derived cancers is caused by silencing ofLARGE
Cellular and Molecular Biology, 2010Co-Authors: Daniel Beltran, Kei-ichiro Inamori, Takako Moriguchi, Christine J. Weydert, Hollie A. Harper, Tobias Willer, William J. Muller, Michael D. Henry, Kevin P. CampbellAbstract:Dystroglycan is a ubiquitously expressed cell membrane receptor that mediates interactions between cells and basement membranes in various epithelia. Dystroglycan is synthesized as a preprotein that is cleaved in two peptides: the transmembranal beta-Dystroglycan, and the extracellular laminin receptor Alpha-Dystroglycan. In many epithelium-derived cancers, beta-Dystroglycan is detected but Alpha-Dystroglycan has been reported to be absent. Here we report that Alpha-Dystroglycan is correctly expressed and trafficked to the cell membrane but lack its laminin-binding capabilities as a result of the silencing of the glycosyltransferase LARGE in a cohort of highly metastatic epithelial cell lines derived from breast, cervical, and lung cancers. As a result of this glycosylation defect, Alpha-Dystroglycan presents a lower MW on Western blot analysis. Exogenous expression of LARGE in these cancer cells restores the normal glycosylation and laminin-binding of Alpha-Dystroglycan leading to enhanced cell adhesion and reduced cell migration in vitro. The relevance of Alpha-Dystroglycan glycosylation in breast cancer is highlighted by the loss of the functionally active form of the receptor during mammary tumor growth in the mouse model for breast cancer MMTV-Neu(YD). Our findings demonstrate that LARGE repression underlies the defective Dystroglycan-mediated cell adhesion that is observed in epithelium-derived cancer cells and uncovers Dystroglycan hypo-glycosylation as factor in cancer progression. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 996.
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o mannosyl phosphorylation of Alpha Dystroglycan is required for laminin binding
Science, 2010Co-Authors: Takako Yoshidamoriguchi, Stephanie H Stalnaker, Sarah Davis, Stefan Kunz, Michael Madson, Michael B A Oldstone, Harry Schachter, Lance Wells, Kevin P. CampbellAbstract:Alpha-Dystroglycan (α-DG) is a cell-surface receptor that anchors the basal lamina to the sarcolemma by binding proteins containing laminin-G domains. This binding is essential for protecting muscle from contraction-induced injury, and defective binding is thought to cause a subclass of congenital muscular dystrophy (CMD) in humans. Mutations in six (putative) glycosyltransferase genes have been identified in patients with CMD, suggesting that glycosylation of α-DG may confer the ability to bind laminin. Despite extensive efforts for over 20 years, the actual laminin-binding moiety has remained unclear. Now, Yoshida-Moriguchi et al. (p. [88][1]) have identified a phosphorylated O -mannosyl glycan on α-DG. This modification occurred in the Golgi via an unidentified kinase and was required for the maturation of α-DG into its laminin-binding form. [1]: /lookup/doi/10.1126/science.1180512
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A comparative study of α-Dystroglycan glycosylation in Dystroglycanopathies suggests that the hypoglycosylation of α-Dystroglycan does not consistently correlate with clinical severity
Brain pathology (Zurich Switzerland), 2008Co-Authors: Cecilia Jimenez-mallebrera, Lucy Feng, Emma Clement, Caroline Godfrey, Silvia Torelli, J. Kim, R. Mein, Stephen Abbs, Susan C. Brown, Kevin P. CampbellAbstract:Hypoglycosylation of Alpha-Dystroglycan underpins a subgroup of muscular dystrophies ranging from congenital onset of weakness, severe brain malformations and death in the perinatal period to mild weakness in adulthood without brain involvement. Mutations in six genes have been identified in a proportion of patients. POMT1, POMT2 and POMGnT1 encode for glycosyltransferases involved in the mannosylation of Alpha-Dystroglycan but the function of fukutin, FKRP and LARGE is less clear. The pathological hallmark is reduced immunolabeling of skeletal muscle with antibodies recognizing glycosylated epitopes on Alpha-Dystroglycan. If the common pathway of these conditions is the hypoglycosyation of Alpha-Dystroglycan, one would expect a correlation between clinical severity and the extent of hypoglycosylation. By studying 24 patients with mutations in these genes, we found a good correlation between reduced Alpha-Dystroglycan staining and clinical course in patients with mutations in POMT1, POMT2 and POMGnT1. However, this was not always the case in patients with defects in fukutin and FKRP, as we identified patients with mild limb-girdle phenotypes without brain involvement with profound depletion of Alpha-Dystroglycan. These data indicate that it is not always possible to correlate clinical course and Alpha-Dystroglycan labeling and suggest that there might be differences in Alpha-Dystroglycan processing in these disorders.