Hypoglycosylation of α-dystroglycan in patients with hereditary IBM due to GNE mutations
Introduction
Hereditary inclusion body myopathy (HIBM; OMIM 600737) is an autosomal recessive neuromuscular disorder characterized by slowly progressive myopathic weakness and atrophy. The disease usually manifests in the second or third decade with foot drop and progressive muscle weakness that involves all limbs but nearly always spares the quadriceps muscles. Histologically, the muscle fibers degenerate and form rimmed vacuoles, especially in atrophic areas [1], [2], [3]. HIBM results from mutations in the UDP-N-acetylglucosamine-2-epimerase/N-acetylmannosamine kinase gene (GNE) on chromosome 9p12–13 [4]. A Japanese variant of HIBM, distal myopathy with rimmed vacuoles (DMRV), is allelic to HIBM and displays mutations in GNE [5]. GNE is the rate limiting, bi-functional enzyme that catalyzes the first two steps committed toward sialic acid biosynthesis [6]. Sialic acid serves as the terminal, charged sugar on glycoproteins and glycolipids, and loss of GNE activity interferes with proper sialylation of cell surface glycoconjugates [7]. However, evidence of impaired sialylation of muscle glycoproteins has not been demonstrated in the muscles of patients with HIBM.
A reasonable candidate for such undersialylation is α-dystroglycan, an essential component of the dystrophin–glycoprotein complex (DGC). Within the DGC, α-dystroglycan binds components of the extracellular matrix, thereby linking it to the actin-associated cytoskeleton via β-dystroglycan and dystrophin [8]. α-Dystroglycan is heavily glycosylated with predominantly O-linked glycans, including rare O-mannosyl linked glycans consisting of ser/thr–mannose–N-acetylglucosamine–galactose–sialic acid. These glycans appear to be critical for interactions with laminin and other extracellular matrix ligands [8], [9]. Abnormal dystroglycan–ligand interactions, some of which occur because of improper glycosylation of α-dystroglycan, provide the explanation for the pathogenesis of several muscular dystrophies, including Fukuyama’s congenital muscular dystrophy (FCMD), Muscle–Eye–Brain disease (MEB), and Walker–Warburg syndrome (WWS) [8], [10]. Perturbation of α-dystroglycan–ligand interactions in muscle fibers, perhaps due to inadequate sialylation of α-dystroglycan as a result of GNE mutations, could explain the clinical manifestations of muscle weakness in HIBM. Here we report markedly reduced α-dystroglycan staining, using glycan-recognizing antibodies, in the skeletal muscle specimens of four HIBM individuals, including two previously unreported patients with novel GNE mutations.
Section snippets
Subjects
Patient #1 is a 39-year-old woman diagnosed at the National Institutes of Health Clinical Center in 1993 as the first non-Iranian Jewish, white American patient with quadriceps-sparing inclusion body myopathy [2]. No consanguinity was present in the family. The initial manifestation was bilateral foot drop at the age of 22 years, followed by progressive, diffuse, limb muscle weakness, and atrophy, except for the quadriceps muscles and abductors of the hips. The same disease began in her older
Results
Mutation analysis on genomic DNA from four HIBM patients revealed compound heterozygous mutations in GNE for all four patients. Each patient has one mutation in the epimerase and one in the kinase domain of GNE. Patient #1 carried V216A and A631V mutations, as reported previously [11]. Patient #2 carried R11W and V696M mutations. Patient #3 carried C303X and V696M; similar mutations have been reported by Eisenberg et al. [12]. Patient #4 had two novel mutations, R202L on exon 2 and G559R on
Discussion
In skeletal muscle membranes, both α- and β-dystroglycan participate in the multimeric dystrophin–glycoprotein complex (DGC). This complex maintains the structural stability of the sarcolemma during cycles of contraction and relaxation by linking cytoskeletal actin (via dystrophin) to components of the extracellular matrix (via α-dystroglycan) [8], [9], [14]. Recent studies have shown that aberrant glycosylation of α-dystroglycan is the underlying biochemical defect in several congenital
Note added in proof
Since submission of the paper, we have repeated the experiments using different (non-commercial) lots of IIH6 and VIA-4 antibodies against a- dystroglycan kindly provided to us by Dr. Campbell. These antibodies do not distinguish a difference between the HIBM patients versus controls. This indicates that there exist differences in epitope recognition between lots of the VIA4 and IIH6 monoclonal antibodies. This observation generates additional interest in the disease and the need to identify
Acknowledgements
The authors greatly appreciate the gift of α-dystroglycan core antibody from Dr. Stephan Kroger.
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