Dysferlin-mediated membrane repair protects the heart from stress-induced left ventricular injury

Abstract

Dilated cardiomyopathy is a life-threatening syndrome that can arise from a myriad of causes, but predisposition toward this malady is inherited in many cases. A number of inherited forms of dilated cardiomyopathy arise from mutations in genes that encode proteins involved in linking the cytoskeleton to the extracellular matrix, and disruption of this link renders the cell membrane more susceptible to injury. Membrane repair is an important cellular mechanism that animal cells have developed to survive membrane disruption. We have previously shown that dysferlin deficiency leads to defective membrane resealing in skeletal muscle and muscle necrosis; however, the function of dysferlin in the heart remains to be determined. Here, we demonstrate that dysferlin is also involved in cardiomyocyte membrane repair and that dysferlin deficiency leads to cardiomyopathy. In particular, stress exercise disturbs left ventricular function in dysferlin-null mice and increases Evans blue dye uptake in dysferlin-deficient cardiomyocytes. Furthermore, a combined deficiency of dystrophin and dysferlin leads to early onset cardiomyopathy. Our results suggest that dysferlin-mediated membrane repair is important for maintaining membrane integrity of cardiomyocytes, particularly under conditions of mechanical stress. Thus, our study establishes what we believe is a novel mechanism underlying the cardiomyopathy that results from a defective membrane repair in the absence of dysferlin. IntroductionDilated cardiomyopathy is the most common type of cardiomyopathy, a condition that can often progress into heart failure and sudden death. Many cases of dilated cardiomyopathy have a genetic etiology. Indeed, inherited forms of idiopathic dilated cardiomyopathy account for at least 30% of dilated cardiomyopathy cases. These are a major cause of severe heart failure and necessitate heart transplantation (1). A number of genes encoding sarcomeric and cytoskeletal proteins have been identified as being responsible for dilated cardiomyopathy: cardiac actin, cardiac troponin T, β-myosin heavy chain, α-tropomyosin, α-actinin, titin, metavinculin, and desmin (reviewed in ref. 2). Mutations in genes such as dystrophin (3, 4), α-sarcoglycan (5), β-sarcoglycan (6), γ-sarcoglycan (7), and δ-sarcoglycan (8, 9), which encode proteins involved in linking the cytoskeleton, sarcolemma, and extracellular matrix, have also been identified as causes of some familial dilated cardiomyopathy. Such mutations lead to the disruption of the cytoskeleton-sarcolemma-extracellular matrix link and thus render the sarcolemma more susceptible to contraction-induced injury (10-12).The plasma membrane provides a physical barrier between the extracellular and intracellular environments, and the maintenance of this barrier is crucial for cell survival; however, plasma-membrane disruption occurs physiologically in certain types of cells,

Basal lamina strengthens cell membrane integrity via the laminin G domain-binding motif of α-dystroglycan

“…In contrast, the apparent rate constants did not differ significantly between the two groups (Large myd , 0.016 Ϯ 0.002 s Ϫ1 ; WT, 0.014 Ϯ 0.003 s Ϫ1 ), suggesting that the membrane repair system is unlikely compromised in Large myd muscle. Consistent with the DG-null muscle having a normal membrane repair system, the FM 1-43 dye did not diffuse into the entire fiber of Large myd muscle as it does in dysferlin-null fibers (31)(32)(33). Also, dysferlin immunostaining on the Large myd muscle was normal (Fig.…”

Membrane damage-induced vesicle–vesicle fusion of dysferlin-containing vesicles in muscle cells requires microtubules and kinesin

“…Dysferlin is a critical component of the membrane repair machinery in both skeletal and cardiac muscle (3,20), and loss of this protein leads to muscular dystrophy (3,31,32). Dysferlincontaining vesicles are thought to play a critical role in muscle resealing, but there have been very few studies attempting to directly examine the behavior of dysferlin-containing vesicles in livemuscle cells in the context of cellular wounding.…”

“…Membrane resealing is a conserved process by which cells are able to survive mechanical disruption of the plasma membrane (14)(15)(16)(17)(18)(19). Dysferlin-null skeletal and cardiac muscle show enhanced uptake of membrane impermeable dye following laser-induced wounding, suggesting that dysferlin may play a role in membrane resealing (3,20). Dysferlin is enriched at sites of cellular damage, and these 'patches' are devoid of plasma membrane proteins, indicating that dysferlin may have been delivered from an intracellular membrane source (3,20).…”

“…Dysferlin-null skeletal and cardiac muscle show enhanced uptake of membrane impermeable dye following laser-induced wounding, suggesting that dysferlin may play a role in membrane resealing (3,20). Dysferlin is enriched at sites of cellular damage, and these 'patches' are devoid of plasma membrane proteins, indicating that dysferlin may have been delivered from an intracellular membrane source (3,20). Electron micrographs of dysferlin-deficient muscle fibers show robust accumulation of vesicles under the sarcolemma, suggesting that in wild-type muscle, dysferlin may play a role in fusion of subsarcolemma vesicles with the plasma membrane and that these vesicles may be critical to repairing the membrane following wounding (3).…”

Genetic Manipulation of Dysferlin Expression in Skeletal Muscle

“…Mice lacking dysferlin (Dysf) exhibit progressive disease in skeletal muscle and cardiac tissue despite having a functional DGC, which is characterized by myofiber necrosis, cycles of degeneration and regeneration, inflammation, and adipocyte replacement. 4,5 Lack of dysferlin also results in the accumulation of vesicles and structural membrane defects as analyzed by electron microscopy, suggesting a role in normal membrane turnover and recycling. 4,6 Disease in dystrophic skeletal muscle is dramatically influenced by the inflammatory response, achieved mainly by infiltration of cytotoxic T-lymphocytes and macrophages.…”