Mikiharu Yoshida

Mutations in the Dystrophin-Associated Protein γ-Sarcoglycan in Chromosome 13 Muscular Dystrophy

Severe childhood autosomal recessive muscular dystrophy (SCARMD) is a progressive muscle-wasting disorder common in North Africa that segregates with microsatellite markers at chromosome 13q12. Here, it is shown that a mutation in the gene encoding the 35-kilodalton dystrophin-associated glycoprotein, γ-sarcoglycan, is likely to be the primary genetic defect in this disorder. The human γ-sarcoglycan gene was mapped to chromosome 13q12, and deletions that alter its reading frame were identified in three families and one of four sporadic cases of SCARMD. These mutations not only affect γ-sarcoglycan but also disrupt the integrity of the entire sarcoglycan complex.

β–sarcoglycan (A3b) mutations cause autosomal recessive muscular dystrophy with loss of the sarcoglycan complex

The dystrophin associated proteins (DAPs) are good candidates for harboring primary mutations in the genetically heterogeneous autosomal recessive muscular dystrophies (ARMD). The transmembrane components of the DAPs can be separated into the dystroglycan and the sarcoglycan complexes. Here we report the isolation of cDNAs encoding the 43 kD sarcoglycan protein β–sarcoglycan (A3b) and the localization of the human gene to chromosome 4q12. We describe a young girl with ARMD with truncating mutations on both alleles. Immunostaining of her muscle biopsy shows specific loss of the components of the sarcoglycan complex β–sarcoglycan, α–sarcoglycan (adhalin), and 35 kD sarcoglycan). Thus secondary destabilization of the sarcoglycan complex may be an important pathophysiological event in ARMD.

From dystrophinopathy to sarcoglycanopathy : Evolution of a concept of muscular dystrophy

Duchenne and Becker muscular dystrophies are collectively termed dystrophinopathy. Dystrophinopathy and severe childhood autosomal recessive muscular dystrophy (SCARMD) are clinically very similar and had not been distinguished in the early 20th century. SCARMD was first classified separately from dystrophinopathy due to differences in the mode of inheritance. Studies performed several years ago clarified some immunohistochemical and genetic characteristics of SCARMD, but many remained to be clarified. In 1994, the sarcoglycan complex was discovered among dystrophin‐associated proteins. Subsequently, on the basis of our immunohistochemical findings which indicated that all components of the sarcoglycan complex are absent in SCARMD muscles, and the previous genetic findings, we proposed that a mutation of any one of the sarcoglycan genes leads to SCARMD. This hypothesis explained and predicted various characteristics of SCARMD at the molecular level, most of which have been verified by subsequent discoveries in our own as well as various other laboratories. SCARMD is now called sarcoglycanopathy, which is caused by a defect of any one of four different sarcoglycan genes, and thus far mutations in sarcoglycan genes have been documented in the SCARMD patients. In this review, the evolution of the concept of sarcoglycanopathy separate from that of dystrophinopathy is explained by comparing studies on these diseases.

Glycoprotein-binding site of dystrophin is confined to the cysteine-rich domain and the first half of the carboxy-terminal domain

Dystrophin, a protein product of the Duchenne muscular dystrophy gene, is thought to associate with the muscle membrane by way of a glycoprotein complex which was co‐purified with dystrophin. Here, we firstly demonstrate direct biochemical evidence for association of the carboxy‐terminal region of dystrophin with the glycoprotein complex. The binding site is found to lie further inward than previously expected and confined to the cysteine‐rich domain and the first half of the carboxy‐terminal domain. Since this portion corresponds well to the region that, when missing results in severe phenotypes, our findings may provide it molecular basis of the disease.

The Three Human Syntrophin Genes Are Expressed in Diverse Tissues, Have Distinct Chromosomal Locations, and Each Bind to Dystrophin and Its Relatives

The syntrophins are a biochemically heterogeneous group of 58-kDa intracellular membrane-associated dystrophin-binding proteins. We have cloned and characterized human acidic (α1-) syntrophin and a second isoform of human basic (β2-) syntrophin. Comparison of the deduced amino acid structure of the three human isoforms of syntrophin (together with the previously reported human β1-syntrophin) demonstrates their overall similarity. The deduced amino acid sequences of human α1- and β2-syntrophin are nearly identical to their homologues in mouse, suggesting a strong functional conservation among the individual isoforms. Much like β1-syntrophin, human β2-syntrophin has multiple transcript classes and is expressed widely, although in a distinct pattern of relative abundance. In contrast, human α1-syntrophin is most abundant in heart and skeletal muscle, and less so in other tissues. Somatic cell hybrids and fluorescent in situ hybridization were both used to determine their chromosomal locations: β2-syntrophin to chromosome 16q22-23 and α1-syntrophin to chromosome 20q11.2. Finally, we used in vitro translated proteins in an immunoprecipitation assay to show that, like β1-syntrophin, both β2- and α1-syntrophin interact with peptides encoding the syntrophin-binding region of dystrophin, utrophin/dystrophin related protein, and the Torpedo 87K protein.

Molecular and cell biology of the sarcoglycan complex

The original sarcoglycan (SG) complex has four subunits and comprises a subcomplex of the dystrophin–dystrophin‐associated protein complex. Each SG gene has been shown to be responsible for limb‐girdle muscular dystrophy, called sarcoglycanopathy (SGP). In this review, we detail the characteristics of the SG subunits, and the mechanism of the formation of the SG complex and various molecules associated with this complex. We discuss the molecular mechanisms of SGP based on studies mostly using SGP animal models. In addition, we describe other SG molecules, ϵ‐ and ζ‐SGs, with special reference to their expression and roles in vascular smooth muscle, which are currently in dispute. We further consider the maternally imprinted nature of the ϵ‐SG gene. Finally, we stress that the SG complex cannot work by itself and works in a larger complex system, called the transverse fixation system, which forms an array of molecules responsible for various muscular dystrophies. Muscle Nerve, 2005

Creatine kinase, cell membrane and Duchenne muscular dystrophy

In 1958 Professor Setsuro Ebashi found that serum creatine kinase activity is increased in patients suffering from various muscular dystrophies, especially Duchenne muscular dystrophy (DMD). He and others proposed that creatine kinase passes through the cell membrane as it is released from DMD muscle fibers.Since then, it has been found that dystrophin and dystrophin-associated proteins are connected to several other components, including the basal lamina and subsarcolemmal cytoskeletal networks on the cell membrane, while dystrophin anchors these dystrophin-associated proteins to the actin filaments inside the muscle cell. In DMD muscle, dystrophin has been found to be absent and dystroglycans and sarcoglycans decreased. However, how creatine kinase molecules can pass through the DMD muscle cell membrane still remains unanswered.On the basis of recent findings on the structure of the protein layers which sandwich the lipid bilayer of muscle cell membranes, this essay stresses the importance of these lipid bilayers in protecting creatine kinase release from protoplasma in normal muscle. It further indicates the possibility that the absence of dystrophin in DMD muscle during muscle contraction may result in temporal damage to the lipid bilayer.

Defective association of dystrophin with sarcolemmal glycoproteins in the cardiomyopathic hamster heart

In ventricular muscle from 30‐ to 60‐day‐old Bio 14.6 cardiomyopathic hamsters, dystrophin‐associated glycoproteins of 43, 50 and 150 kDa are markedly reduced in abundance. In particular, the 50‐kDa glycoprotein is totally deficient in the sareolemma of myopathic ventricular myocytes as revealed by immunofluorescence microscopy. The dystrophin‐glycoprotein complex formation is defective in the cardiomyopathic hamster heart, because dystrophin and the glycoproteins behave independently when digitonin‐solubilized ventricular homogenates are fractionated on wheat germ agglutinin beads or anti‐dystrophin immunoaffinity beads.

Dystrophin-associated protein A0 is a homologue of the Torpedo 87K protein

We raised a monoclonal antibody, MA0, which reacts with A0, a 94‐kDa rabbit skeletal muscle dystrophin‐associated protein (DAP) bound to the syntrophin‐binding domain of dystrophin. The antibody also reacted with the 62‐kDa DAP which was moved to the locus close to β‐syntrophins by 2‐dimensional PAGE, but the DAP did not coincide with any known β‐syntrophins. We have cloned a fragment of cDNA which codes the protein reacting with MA0 from a neonatal rabbit heart cDNA library. Based on the coincidence of cDNA sequences and the similarity in molecular mass, we concluded that the proteins reacting with MA0 are rabbit homologues of the Torpedo 87K protein.

THE FOURTH COMPONENT OF THE SARCOGLYCAN COMPLEX

© 1997 Federation of European Biochemical Societies.