A new monoclonal antibody DAG-6F4 against human alpha-dystroglycan reveals reduced core protein in some, but not all, dystroglycanopathy patients.

Abstract

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.

• Authors: Emma L Humphrey, Erica Lacey, Lam T Le, Lucy Feng, Francesca Sciandra, Charlotte R Morris, Jane E Hewitt, Ian Holt, Andrea Brancaccio, Rita Barresi, Caroline A Sewry, Susan C Brown, Glenn E Morris
• Journal: Neuromuscular disorders : NMD (Neuromuscul Disord) Vol. 25 Issue 1 Pg. 32-42 (Jan 2015) ISSN: 1873-2364 [Electronic] England
• PMID: 25387694 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2014 Elsevier B.V. All rights reserved.

Chemical References

• Antibodies, Monoclonal
• DAG-6F4 monoclonal antibody
• Membrane Proteins
• Dystroglycans

Topics

• Adult
• Amino Acid Sequence
• Animals
• Antibodies, Monoclonal (immunology)
• Child, Preschool
• Dystroglycans (analysis, metabolism)
• Glycosylation
• HEK293 Cells
• Humans
• Immunohistochemistry
• Infant
• Membrane Proteins (immunology)
• Mice
• Mice, Inbred BALB C
• Molecular Sequence Data
• Muscle, Skeletal (immunology)
• Muscular Dystrophy, Duchenne (diagnosis, pathology)
• Sarcolemma (immunology)