Myofibrillar myopathy (MFM) is a human disease that is characterized by focal myofibrillar destruction and pathological cytoplasmic protein aggregations. In an extended German pedigree with a novel form of MFM characterized by clinical features of a limb-girdle myopathy and morphological features of MFM, we identified a co-segregating, heterozygous nonsense mutation (8130G-->A; W2710X) in the filamin c gene (FLNC) on chromosome 7q32.1. The mutation is the first found in FLNC and is localized in the dimerization domain of filamin c. Functional studies showed that, in the truncated mutant protein, this domain has a disturbed secondary structure that leads to the inability to dimerize properly. As a consequence of this malfunction, the muscle fibers of our patients display massive cytoplasmic aggregates containing filamin c and several Z-disk-associated and sarcolemmal proteins.

SummaryThe UCS (UNC-45/CRO1/She4) chaperones play an evolutionarily conserved role in promoting myosin-dependent processes, including cytokinesis, endocytosis, RNA transport, and muscle development. To investigate the protein machinery orchestrating myosin folding and assembly, we performed a comprehensive analysis of Caenorhabditis elegans UNC-45. Our structural and biochemical data demonstrate that UNC-45 forms linear protein chains that offer multiple binding sites for cooperating chaperones and client proteins. Accordingly, Hsp70 and Hsp90, which bind to the TPR domain of UNC-45, could act in concert and with defined periodicity on captured myosin molecules. In vivo analyses reveal the elongated canyon of the UCS domain as a myosin-binding site and show that multimeric UNC-45 chains support organization of sarcomeric repeats. In fact, expression of transgenes blocking UNC-45 chain formation induces dominant-negative defects in the sarcomere structure and function of wild-type worms. Together, these findings uncover a filament assembly factor that directly couples myosin folding with myofilament formation.

Xin is a protein that is expressed during early developmental stages of cardiac and skeletal muscles. Immunolocalization studies indicated a peripheral localization in embryonic mouse heart, where Xin localizes with β-catenin and N-cadherin. In adult tissues, Xin is found primarily in the intercalated discs of cardiomyocytes and the myotendinous junctions of skeletal muscle cells, both specialized attachment sites of the myofibrillar ends to the sarcolemma. A large part of the Xin protein consists of unique 16 amino acid repeats with unknown function. We have investigated the characteristics of the Xin repeats by transfection experiments and actin-binding assays and ascertained that, upon expression in cultured cells, these repeats bind to and stabilize the actin-based cytoskeleton. In vitro co-sedimentation assays with skeletal muscle actin indicated that they not only directly bind actin filaments, but also have the capability of arranging microfilaments into networks that sediment upon low-speed centrifugation. Very similar repeats were also found in `Xin-repeat protein 2' (XIRP2), a novel protein that seems to be expressed mainly in striated muscles. Human XIRP2 contains 28 Xin repeats with properties identical to those of Xin. We conclude that the Xin repeats define a novel, repetitive actin-binding motif present in at least two different muscle proteins. These Xin-repeat proteins therefore constitute the first two members of a novel family of actin-binding proteins.

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