The 16 kDa subunit of vacuolar H+-ATPase is a novel sarcoglycan-interacting protein

The sarcoglycan complex in muscle consists of alpha-, beta-, gamma- and delta-sarcoglycan and is part of the larger dystrophin-glycoprotein complex (DGC), which is essential for maintaining muscle membrane integrity. Mutations in any of the four sarcoglycans cause limb-girdle muscular dystrophies (LGMD). In this report, we have identified a novel interaction between delta-sarcoglycan and the 16 kDa subunit c (16K) of vacuolar H(+)-ATPase. Co-expression studies in heterologous cell system revealed that 16K interacts specifically with delta-sarcoglycan and the highly related gamma-sarcoglycan through the transmembrane domains. In cultured C2C12 myotubes, 16K forms a complex with sarcoglycans at the plasma membrane. Loss of sarcoglycans in the sarcoglycan-deficient BIO14.6 hamster destabilizes the DGC and alters the localization of 16K at the sarcolemma. In addition, the steady state level of beta(1)-integrin is increased. Recent studies have shown that 16K also interacts directly with beta(1)-integrin and our data demonstrated that sarcoglycans, 16K and beta(1)-integrin were immunoprecipitated together in C2C12 myotubes. Since sarcoglycans have been proposed to participate in bi-directional signaling with integrins, our findings suggest that 16K might mediate the communication between sarcoglycans and integrins and play an important role in the pathogenesis of muscular dystrophy.     

Filamin 2 (FLN2): A muscle-specific sarcoglycan interacting protein

Mutations in genes encoding for the sarcoglycans, a subset of proteins within the dystrophin-glycoprotein complex, produce a limb-girdle muscular dystrophy phenotype; however, the precise role of this group of proteins in the skeletal muscle is not known. To understand the role of the sarcoglycan complex, we looked for sarcoglycan interacting proteins with the hope of finding novel members of the dystrophin-glycoprotein complex. Using the yeast two-hybrid method, we have identified a skeletal muscle-specific form of filamin, which we term filamin 2 (FLN2), as a gamma- and delta-sarcoglycan interacting protein. In addition, we demonstrate that FLN2 protein localization in limb-girdle muscular dystrophy and Duchenne muscular dystrophy patients and mice is altered when compared with unaffected individuals. Previous studies of filamin family members have determined that these proteins are involved in actin reorganization and signal transduction cascades associated with cell migration, adhesion, differentiation, force transduction, and survival. Specifically, filamin proteins have been found essential in maintaining membrane integrity during force application. The finding that FLN2 interacts with the sarcoglycans introduces new implications for the pathogenesis of muscular dystrophy.     

The 16 kDa subunit of vacuolar H+-ATPase is a novel sarcoglycan-interacting protein

The sarcoglycan complex in muscle consists of α-, β-, γ- and δ-sarcoglycan and is part of the larger dystrophin–glycoprotein complex (DGC), which is essential for maintaining muscle membrane integrity. Mutations in any of the four sarcoglycans cause limb-girdle muscular dystrophies (LGMD). In this report, we have identified a novel interaction between δ-sarcoglycan and the 16 kDa subunit c (16K) of vacuolar H⁺-ATPase. Co-expression studies in heterologous cell system revealed that 16K interacts specifically with δ-sarcoglycan and the highly related γ-sarcoglycan through the transmembrane domains. In cultured C2C12 myotubes, 16K forms a complex with sarcoglycans at the plasma membrane. Loss of sarcoglycans in the sarcoglycan-deficient BIO14.6 hamster destabilizes the DGC and alters the localization of 16K at the sarcolemma. In addition, the steady state level of β₁-integrin is increased. Recent studies have shown that 16K also interacts directly with β₁-integrin and our data demonstrated that sarcoglycans, 16K and β₁-integrin were immunoprecipitated together in C2C12 myotubes. Since sarcoglycans have been proposed to participate in bi-directional signaling with integrins, our findings suggest that 16K might mediate the communication between sarcoglycans and integrins and play an important role in the pathogenesis of muscular dystrophy.     

Molecular Organization of Sarcoglycan Complex in Mouse Myotubes in Culture

The sarcoglycans are a complex of four transmembrane proteins (α, β, γ, and δ) which are primarily expressed in skeletal muscle and are closely associated with dystrophin and the dystroglycans in the muscle membrane. Mutations in the sarcoglycans are responsible for four autosomal recessive forms of muscular dystrophy. The function and the organization of the sarcoglycan complex are unknown. We have used coimmunoprecipitation and in vivo cross-linking techniques to analyze the sarcoglycan complex in cultured mouse myotubes. We demonstrate that the interaction between β- and δ-sarcoglycan is resistant to high concentrations of SDS and α-sarcoglycan is less tightly associated with other members of the complex. Cross-linking experiments show that β-, γ-, and δ-sarcoglycan are in close proximity to one another and that δ-sarcoglycan can be cross-linked to the dystroglycan complex. In addition, three of the sarcoglycans (β, γ, and δ) are shown to form intramolecular disulfide bonds. These studies further our knowledge of the structure of the sarcoglycan complex. Our proposed model of their interactions helps to explain some of the emerging data on the consequences of mutations in the individual sarcoglycans, their effect on the complex, and potentially the clinical course of muscular dystrophies.     

Dynamic changes in the subcellular localization of Drosophila beta-sarcoglycan during the cell cycle

One of the proposed roles of sarcoglycan is to stabilize dystrophin glycoprotein complexes in muscle sarcolemma. Involvement in signal transduction has also been proposed and abnormalities in some sarcoglycan genes are known to be responsible for muscular dystrophy. While characterization of sarcoglycans in muscle has been performed, little is known about its functions in the non-muscle tissues in which mammalian sarcoglycans are expressed. Here, we investigated temporal and spatial expression patterns of Drosophila beta-sarcoglycan (dScgbeta) during development by immunohistochemistry. In addition to almost ubiquitous expression in various tissues and organs, as seen for its mammalian counterpart, anti-dScgbeta staining data of embryos, eye imaginal discs, and salivary glands demonstrated cytoplasmic localization during S phase in addition to plasma membrane staining. Furthermore we found that subcellular localization of dScgbeta dramatically changes during mitosis through possible association with tubulin. These observations point to a complex role of sarcoglycans in non-muscle tissues.     

Sarcoglycan Complex in Masseter and Sternocleidomastoid Muscles of Baboons: An Immunohistochemical Study

The sarcoglycan complex consists of a group of single-pass transmembrane glycoproteins that are essential to maintain the integrity of muscle membranes. Any mutation in each sarcoglycan gene causes a series of recessive autosomal dystrophin-positive muscular dystrophies. Negative fibres for sarcoglycans have never been found in healthy humans and animals. In this study, we have investigated whether the social ranking has an influence on the expression of sarcoglycans in the skeletal muscles of healthy baboons. Biopsies of masseter and sternocleidomastoid muscles were processed for confocal immunohistochemical detection of sarcoglycans. Our findings showed that baboons from different social rankings exhibited different sarcoglycan expression profiles. While in dominant baboons almost all muscles were stained for sarcoglycans, only 55% of muscle fibres showed a significant staining. This different expression pattern is likely to be due to the living conditions of these primates. Sarcoglycans which play a key role in muscle activity by controlling contractile forces may influence the phenotype of muscle fibres, thus determining an adaptation to functional conditions. We hypothesize that this intraspecies variation reflects an epigenetic modification of the muscular protein network that allows baboons to adapt progressively to a different social status.Key words: Sarcoglycan, baboons, immunohistochemistry, evolution, ranking     

Functional characteristics of dystrophic skeletal muscle: insights from animal models.

Muscular dystrophies are a clinically and genetically heterogeneous group of disorders that show myofiber degeneration and regeneration. Identification of animal models of muscular dystrophy has been instrumental in research on the pathogenesis, pathophysiology, and treatment of these disorders. We review our understanding of the functional status of dystrophic skeletal muscle from selected animal models with a focus on 1) the mdx mouse model of Duchenne muscular dystrophy, 2) the Bio 14.6 delta-sarcoglycan-deficient hamster model of limb-girdle muscular dystrophy, and 3) transgenic null mutant murine lines of sarcoglycan (alpha, beta, delta, and gamma) deficiencies. Although biochemical data from these models suggest that the dystrophin-sarcoglycan-dystroglycan-laminin network is critical for structural integrity of the myofiber plasma membrane, emerging studies of muscle physiology suggest a more complex picture, with specific functional deficits varying considerably from muscle to muscle and model to model. It is likely that changes in muscle structure and function, downstream of the specific, primary biochemical deficiency, may alter muscle contractile properties.     

Sarcoglycan isoforms in skeletal muscle

The heterotetrameric sarcoglycan complex, composed of alpha-, beta-, gamma-, and delta-sarcoglycans, is an important component of the dystrophin-associated glycoprotein assembly in striated muscle. Mutations in any of the four genes encoding sarcoglycans cause a deficiency in all sarcoglycans in the sarcolemma and produce one of four types of limb-girdle muscular dystrophy. A fifth widely expressed sarcoglycan, epsilon-sarcoglycan, has been recently described. epsilon-Sarcoglycan is homologous to alpha-sarcoglycan, but whether it associates with the other sarcoglycans in muscle is not known. In this study, we use wild type and alpha-sarcoglycan-deficient mice to analyze the localization and association of sarcoglycans in skeletal muscle in vivo. The amounts of beta-, gamma-, and delta-sarcoglycans are reduced in alpha-sarcoglycan mutants, whereas the amount of epsilon-sarcoglycan is unchanged. We show here that epsilon-sarcoglycan is complexed with beta-, gamma-, and delta-sarcoglycans in both wild type and alpha-sarcoglycan mutant mice. We also use C2C12 myocytes to study the temporal expression and organization of sarcoglycan complexes during muscle cell differentiation in vitro. In C2C12 cells, alpha- and epsilon-sarcoglycans form separate complexes with beta-, gamma-, and delta-sarcoglycans. Both types of complexes are expressed at the cell surface and presumed to be functional. These results suggest that epsilon-sarcoglycan serves a function similar to that of alpha-sarcoglycan and that residual beta-, gamma-, and delta-sarcoglycan seen in mutant mice and alpha-sarcoglycan-deficient patients is due to its association with epsilon-sarcoglycan.     

Stable restoration of the sarcoglycan complex in dystrophic muscle perfused with histamine and a recombinant adeno-associated viral vector

Limb-girdle muscular dystrophies 2C-F represent a family of autosomal recessive diseases caused by defects in sarcoglycan genes. The cardiomyopathic hamster is a naturally occurring model for limb-girdle muscular dystrophy caused by a primary deficiency in delta-sarcoglycan. We show here that acute sarcolemmal disruption occurs in this animal model during forceful muscle contraction. A recombinant adeno-associated virus vector encoding human delta-sarcoglycan conferred efficient and stable genetic reconstitution in the adult cardiomyopathic hamster when injected directly into muscle. A quantitative assay demonstrated that vector-transduced muscle fibers are stably protected from sarcolemmal disruption; there was no associated inflammation or immunologic response to the vector-encoded protein. Efficient gene transduction with rescue of the sarcoglycan complex in muscle fibers of the distal hindlimb was also obtained after infusion of recombinant adeno-associated virus into the femoral artery in conjunction with histamine-induced endothelial permeabilization. This study provides a strong rationale for the development of gene therapy for limb-girdle muscular dystrophy.     

epsilon-sarcoglycan replaces alpha-sarcoglycan in smooth muscle to form a unique dystrophin-glycoprotein complex.

The sarcoglycan complex has been well characterized in striated muscle, and defects in its components are associated with muscular dystrophy and cardiomyopathy. Here, we have characterized the smooth muscle sarcoglycan complex. By examination of embryonic muscle lineages and biochemical fractionation studies, we demonstrated that epsilon-sarcoglycan is an integral component of the smooth muscle sarcoglycan complex along with beta- and delta-sarcoglycan. Analysis of genetically defined animal models for muscular dystrophy supported this conclusion. The delta-sarcoglycan-deficient cardiomyopathic hamster and mice deficient in both dystrophin and utrophin showed loss of the smooth muscle sarcoglycan complex, whereas the complex was unaffected in alpha-sarcoglycan null mice in agreement with the finding that alpha-sarcoglycan is not expressed in smooth muscle cells. In the cardiomyopathic hamster, the smooth muscle sarcoglycan complex, containing epsilon-sarcoglycan, was fully restored following intramuscular injection of recombinant delta-sarcoglycan adenovirus. Together, these results demonstrate a tissue-dependent variation in the sarcoglycan complex and show that epsilon-sarcoglycan replaces alpha-sarcoglycan as an integral component of the smooth muscle dystrophin-glycoprotein complex. Our results also suggest a molecular basis for possible differential smooth muscle dysfunction in sarcoglycan-deficient patients.     

Alpha-sarcoglycan is recycled from the plasma membrane in the absence of sarcoglycan complex assembly

The sarcoglycan complex consists of four subunits in skeletal muscle (alpha, beta, gamma, and delta-SG). Mutations in alpha-sarcoglycan (alpha-SG) result in the most common form of limb girdle muscular dystrophy. However, the function of alpha-SG remains unknown. In this report we attempt to clarify its function by delineating the trafficking pathway of alpha-SG in live cells. We present evidence, utilizing total internal reflection microscopy, fluorescence recovery after photobleaching and photoactivation of green fluorescent protein (GFP) constructs, that pools of alpha-SG are able to translocate to the plasma membrane in the absence of the remaining sarcoglycans. Internalization assays and drug treatment experiments demonstrate that alpha-SG recycles from the plasma membrane and accumulates in recycling endosomes. We also establish that alpha-SG utilizes well-described clathrin mediated mechanisms and microtubules to traffic within the cell. Finally, we show that the most commonly reoccurring limb girdle muscular dystrophy (R77C) mutation causes a fundamental defect in protein biosynthesis, trapping the mutant protein in the endoplasmic recticulum (ER). These results demonstrate that alpha-SG requires assembly into the sarcoglycan complex for stability at the plasma membrane rather than export out of the ER. Furthermore, this data suggests that alpha-SG utilizes known trafficking machinery to control deposition at the plasma membrane through recycling.     

Isolation and characterization of myogenic satellite cells from the muscular dystrophic hamster

Myogenic satellite cells were isolated from control and dystrophic hamster diaphragms to examine cellular mechanisms involved in the physiology of muscular dystrophy. The Bio 14.6 dystrophic hamster, which possesses a defect in the delta-sarcoglycan gene, develops biochemical and physical symptoms of Duchenne-like and limb girdle muscular dystrophies. Because primary cultures of the control and dystrophic satellite cells became extensively contaminated with non-myogenic cells during proliferation, cell clones were developed to provide pure cultures for study. Cell culture conditions were optimized with the use of Ham's F-12K medium containing 10% fetal bovine serum +5% horse serum + 10 ng/mL basic fibroblast growth factor + 50 microg/mL porcine gelatin. Proliferation rates of the two clonal cultures were similar between the two lines. Satellite cell-derived myotubes from both primary cultures and clones differed between control and dystrophic animals. Dystrophic myotubes tended to be long and narrow, while the control-derived myotubes were broader. Measurement of muscle-specific creatine kinase during differentiation revealed that the dystrophic myotubes possessed higher creatine kinase levels than control myotubes (up to 146-fold at 168 h). The results demonstrate that satellite cells can be isolated from the hamster and may provide a useful tool to study muscular dystrophies associated with defects in the sarcoglycan complex and the involvement of sarcoglycans in normal skeletal muscle growth and development.     

Delta-sarcoglycan is necessary for early heart and muscle development in zebrafish

Delta-sarcoglycan, one member of the sarcoglycan complex, is a very conservative muscle-specific protein exclusively expressed in the skeletal and cardiac muscles of vertebrates. Mutations in sarcoglycans are known to be involved in limb-girdle muscular dystrophy (LGMD) and dilated cardiomyopathy (DCM) in humans. To address the role of delta-sarcoglycan gene in zebrafish development, we have studied expression pattern of delta-sarcoglycan in zebrafish embryos and examined the role of delta-sarcoglycan in zebrafish embryonic development by morpholino. Strong expression of delta-sarcoglycan was observed in various muscles including those of the segment, heart, eye, jaw, pectoral fin, branchial arches, and swim bladder in zebrafish embryo. Delta-sarcoglycan was also expressed in midbrain and retina. Knockdown of delta-sarcoglycan resulted in severe abnormality in both the cardiac and skeletal muscles. Some severe ones displayed serious morphological abnormality such as hypoplastic head, linear heart, very weak heartbeats, and runtish trunk, all dead within 5 dpf. Whole-mount in situ hybridization analysis showed that adaxial cells and muscle pioneers were affected in delta-sarcoglycan knockdown embryos. In addition, absence of delta-sarcoglycan protein severely delayed the cardiac development and influenced the differentiation of cardiac muscle, and the cardiac left-right asymmetry was dramatically changed in morpholino-treated embryos. These data together suggest that delta-sarcoglycan plays an important role in early heart and muscle development.     

Overexpression of gamma-sarcoglycan induces severe muscular dystrophy. Implications for the regulation of Sarcoglycan assembly

The sarcoglycan complex is found normally at the plasma membrane of muscle. Disruption of the sarcoglycan complex, through primary gene mutations in dystrophin or sarcoglycan subunits, produces membrane instability and muscular dystrophy. Restoration of the sarcoglycan complex at the plasma membrane requires reintroduction of the mutant sarcoglycan subunit in a manner that will permit normal assembly of the entire sarcoglycan complex. To study sarcoglycan gene replacement, we introduced transgenes expressing murine gamma-sarcoglycan into muscle of normal mice. Mice expressing high levels of gamma-sarcoglycan, under the control of the muscle-specific creatine kinase promoter, developed a severe muscular dystrophy with greatly reduced muscle mass and early lethality. Marked gamma-sarcoglycan overexpression produced cytoplasmic aggregates that interfered with normal membrane targeting of gamma-sarcoglycan. Overexpression of gamma-sarcoglycan lead to the up-regulation of alpha- and beta-sarcoglycan. These data suggest that increased gamma-sarcoglycan and/or mislocalization of gamma-sarcoglycan to the cytoplasm is sufficient to induce muscle damage and provides a new model of muscular dystrophy that highlights the importance of this protein in the assembly, function, and downstream signaling of the sarcoglycan complex. Most importantly, gene dosage and promoter strength should be given serious consideration in replacement gene therapy to ensure safety in human clinical trials.     

Biglycan binds to alpha- and gamma-sarcoglycan and regulates their expression during development

The dystrophin-associated protein complex (DAPC), which links the cytoskeleton to the extracellular matrix, is essential for muscle cell survival, and is defective in a wide range of muscular dystrophies. The DAPC contains two transmembrane subcomplexes-the dystroglycans and the sarcoglycans. Although several extracellular binding partners have been identified for the dystroglycans, none have been described for the sarcoglycan subcomplex. Here we show that the small leucine-rich repeat (LRR) proteoglycan biglycan binds to alpha- and gamma-sarcoglycan as judged by ligand blot overlay and co-immunoprecipitation assays. Our studies with biglycan-decorin chimeras show that alpha- and gamma-sarcoglycan bind to distinct sites on the polypeptide core of biglycan. Both biglycan proteoglycan as well as biglycan polypeptide lacking glycosaminoglycan (GAG) side chains are components of the dystrophin glycoprotein complex isolated from adult skeletal muscle membranes. Finally, our immunohistochemical and biochemical studies with biglycan null mice show that the expression of alpha- and gamma-sarcoglycan is selectively reduced in muscle from young (P14-P21) animals, while levels in adult muscle (> or = P35) are unchanged. We conclude that biglycan is a ligand for two members of the sarcoglycan complex and regulates their expression at discrete developmental ages.     

Ecto-ATPase activity of alpha-sarcoglycan (adhalin)

alpha-Sarcoglycan is a component of the sarcoglycan complex of dystrophin-associated proteins. Mutations of any of the sarcoglycan genes cause specific forms of muscular dystrophies, collectively termed sarcoglycanopathies. Importantly, a deficiency of any specific sarcoglycan affects the expression of the others. Thus, it appears that the lack of sarcoglycans deprives the muscle cell of an essential, yet unknown function. In the present study, we provide evidence for an ecto-ATPase activity of alpha-sarcoglycan. alpha-Sarcoglycan binds ATP in a Mg2+-dependent and Ca2+-independent manner. The binding is inhibited by 3'-O-(4-benzoyl)benzoyl ATP and ADP. Sequence analysis reveals the existence of a consensus site for nucleotide binding in the extracellular domain of the protein. An antibody against this sequence inhibits the binding of ATP. A dystrophin.dystrophin-associated protein preparation demonstrates a Mg-ATPase activity that is inhibited by the antibody but not by inhibitors of endo-ATPases. In addition, we demonstrate the presence in the sarcolemmal membrane of a P2X-type purinergic receptor. These data suggest that alpha-sarcoglycan may modulate the activity of P2X receptors by buffering the extracellular ATP concentration. The absence of alpha-sarcoglycan in sarcoglycanopathies leaves elevated the concentration of extracellular ATP and the persistent activation of P2X receptors, leading to intracellular Ca2+ overload and muscle fiber death.     

delta- and gamma-Sarcoglycan localization in the sarcoplasmic reticulum of skeletal muscle.

Sarcoglycans are transmembrane proteins that are members of the dystrophin complex. Sarcoglycans cluster together to form a complex, which is localized in the cell membrane of skeletal, cardiac, and smooth muscle fibers. However, it is still unclear whether or not sarcoglycans are restricted to the sarcolemma. To address this issue, we examined alpha-, beta-, delta-, and gamma-sarcoglycan expression in femoral skeletal muscle from control and dystrophin-deficient mice and rats using confocal microscopy and immunoelectron microscopy. Confocal microscopy of the tissues in cross-section showed that all sarcoglycans were detected under the sarcolemma in rats and control mice. delta- and gamma-sarcoglycan labeling demonstrated striations in the longitudinal section, suggesting that the proteins were expressed in the sarcoplasmic reticulum (SR) or transverse tubules (T-tubules). Moreover, such striations of both sarcoglycans were recognized in the dystrophin-deficient mouse skeletal muscle. Double labeling with phalloidin or alpha-actinin and delta- or gamma-sarcoglycan showed different labeling patterns, indicating that delta-sarcoglycan localization was distinct from that of gamma-sarcoglycan. Immunoelectron microscopy clarified that delta-sarcoglycan was localized in the terminal cisternae of the SR, while gamma-sarcoglycan was found in the terminal cisternae and longitudinal SR over I-bands but not over A-bands. These data demonstrate that delta- and gamma-sarcoglycans are components of the SR in skeletal muscle, suggesting that both sarcoglycans function independent of the dystrophin complex in the SR.     

Sarcoglycan subcomplex in normal and pathological human muscle fibers

Sarcoglycans are a sub-complex of transmembrane proteins which are part of the dystrophin-glycoprotein complex (DGC). They are expressed above all in the skeletal, cardiac and smooth muscle. Although numerous studies have been conducted on the sarcoglycan sub-complex in skeletal and cardiac muscle, the manner of distribution and localization of these proteins along the non-junctional sarcolemma is still not clear. Furthermore, there are unclear data about the actual role of sarcoglycans in human skeletal muscle affected by sarcoglycanopathies. In our studies on human skeletal muscle, normal and pathological, we determined the localization, distribution and interaction of these glycoproteins. Our results, on normal human skeletal muscle, showed that the sarcoglycans can be localized both in the region of the sarcolemma over the I band and over the A band, hypothesizing a correlation between regions of the sarcolemma occupied by costameres and the metabolic type of the fibers (slow and fast). Our data on skeletal muscle affected by sarcoglycanopathy confirmed the hypothesis of a bidirectional signaling between sarcoglycans and integrins and the interaction of filamin2 with both sarcoglycans and integrins. In addition, we have recently demonstrated, in smooth muscle, the presence of alpha-SG, in contrast with data of other Authors. Finally, we analyzed the association between contractile activity and quantitative correlation between alpha- and epsilon-SG, in order to better define the arrangement of sarcoglycan subcomplex.     

Ultrastructure of diaphragm from dystrophic alpha-sarcoglycan-null mice

alpha-Sarcoglycan is a 50 kDa single-pass transmembrane glycoprotein exclusively expressed in striated muscle that, together with beta-, gamma-, and delta-sarcoglycan, forms a sub-complex at the muscle fibre cell membrane. The sarcoglycans are components of the dystrophin-associated glycoprotein (DAG) complex which forms a mechanical link between the intracellular cytoskeleton and extracellular matrix. The DAG complex function is to protect the muscle membrane from the stress of contractile activity and as a structure for the docking of signalling proteins. Genetic defects of DAG components cause muscular dystrophies. A lack or defects of alpha-sarcoglycan causes the severe type 2D limb girdle muscular dystrophy. alpha-Sarcoglycan-null (Sgca-null) mice develop progressive muscular dystrophy similar to the human disorder. This animal model was used in the present work for an ultrastructural study of diaphragm muscle. Diaphragm from Sgca-null mouse presents a clear dystrophic phenotype, with necrosis, regeneration, fibre hypertrophy and splitting, excess of collagen and fatty infiltration. Some abnormalities were also observed, such as centrally located nuclei of abnormal shape, fibres containing inclusion bodies within the contractile structure, and fibres with electron-dense material dispersed over almost the entire cell. Additionally, unusual interstitial cells of uncertain identity were detected within muscle fibres. The abnormal ultrastructure of the diaphragm from Sgca-null mice is discussed.