FLNC myofibrillar myopathy results from impaired autophagy and protein insufficiency

Abstract

Myofibrillar myopathy is a progressive muscle disease characterized by the disintegration of muscle fibers and formation of protein aggregates. Causative mutations have been identified in nine genes encoding Z-disk proteins, including the actin binding protein filamin C (FLNC). To investigate the mechanism of disease in FLNC myopathy we overexpressed fluorescently tagged FLNC or FLNC in zebrafish. Expression of FLNC causes formation of protein aggregates but surprisingly, our studies reveal that the mutant protein localizes correctly to the Z-disk and is capable of rescuing the fiber disintegration phenotype that results from FLNC knockdown. This demonstrates that the functions necessary for muscle integrity are not impaired, and suggests that it is the formation of protein aggregates and subsequent sequestration of FLNC away from the Z-disk that results in myofibrillar disintegration. Similar to those found in patients, the aggregates in FLNC expressing fish contain the co-chaperone BAG3. FLNC is a target of the BAG3-mediated chaperone assisted selective autophagy (CASA) pathway and therefore we investigated its role, and the role of autophagy in general, in clearing protein aggregates. We reveal that despite BAG3 recruitment to the aggregates they are not degraded via CASA. Additionally, recruitment of BAG3 is sufficient to block alternative autophagy pathways which would otherwise clear the aggregates. This blockage can be relieved by reducing BAG3 levels or by stimulating autophagy. This study therefore identifies both BAG3 reduction and autophagy promotion as potential therapies for FLNC myofibrillar myopathy, and identifies protein insufficiency due to sequestration, compounded by impaired autophagy, as the cause.

Filamin actin-binding and titin-binding fulfill distinct functions in Z-disc cohesion

“…Like FLNc in vertebrates all Cher isoforms localize to the Z-disc [46,47]. This can now be explained by the titin interaction domain, which we mapped to Cher Ig 19–22, and which is present in all isoforms.…”

“…The phenotypes of these mutations fall in several different classes, and it is often unclear how the phenotype is caused at a cell biological level [29,45]. Recently, a FLNc mutant in zebrafish was generated revealing a mild muscle fiber phenotype [46], confirming the role of filamin in vertebrate muscles.…”

Drosophila vinculin is more harmful when hyperactive than absent, and can circumvent integrin to form adhesion complexes

“…Cytoskeletal machinery is crucial for muscle fusion and muscle pathfinding to tendon cell targets (Maartens and Brown, 2015b), and an interaction between hyperactive vinculin complexes and more general cytoskeletal factors might explain the muscle defects. Sequestration of Z-disc proteins to ectopic intracellular aggregates has been implicated in the muscle phenotypes associated with myofibrillar myopathy (Ruparelia et al, 2016), and a similar effect may be stimulated by hyperactive vinculin.…”

New Developments in the Genetics of Inclusion Body Myositis

“…This candidate-based approach uncovered an increased frequency of rare variants in multiple genes including VCP, SQSTM1, FLNC, ZASP, and BAG3 as well as a novel variant in HNRPA2B1. FLNC, SQSTM1, ZASP, and BAG3 are known to be mutated in rare inherited vacuolar myopathies and also play integral roles in the autophagy pathway [36–38]. SQSTM1 (Sequestosome 1, also known as p62) is a ubiquitin-binding autophagy adaptor that has been shown to label protein inclusions in IBM muscle [39–41].…”