Dysregulated calcium homeostasis prevents plasma membrane repair in Anoctamin 5/TMEM16E-deficient patient muscle cells

Chandra, Goutam; Defour, Aurélia; Mamchoui, Kamel; Pandey, Kalpana; Mishra, Soumya Ranjan; Mouly, Vincent; Sreetama, SenChandra; Ahmad, Mohammad Mahad; Mahjneh, Ibrahim; Morizono, Hiroki; Pattabiraman, Nagarajan; Menon, Anant K.; Jaiswal, Jyoti K.

doi:10.1038/s41420-019-0197-z

Cited by 33 publications

(58 citation statements)

References 59 publications

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“…Our results provide additional support for the growing literature implicating ANO5 in muscle membrane repair while providing novel mechanistic insights about this process. Consistent with previous results (8,52), we show that ANO5-KO myofibers are more permeant to FM1-43 dye after injury. It is important to note that the FM1-43 assay, by itself, is incapable of resolving whether fibers are defective in repair or more susceptible to injury.…”

Section: Discussionsupporting

confidence: 93%

“…We electroporated ANO5-R58W-mCherry into ANO5-KO fibers and found that, at least under condition of overexpression, it was well expressed (Fig. S4), in contrast to the previously reported R758C patient mutation, which leads to ANO5 degradation (8). Expression of ANO5-R58W in ANO5-KO fibers had no effect on FM1-43 accumulation or patch volumes ( Fig.…”

Section: Introductionmentioning

confidence: 77%

“…One suggestion is that ANO5 myopathies are caused by defects in the ability of muscle fibers to self-repair (1,8,24,25). Skeletal muscle endures considerable mechanical stress even during normal use, which can produce small tears in the plasma membrane (sarcolemma) (26).…”

Section: Significance Statementmentioning

confidence: 99%

“…Anoctamin-5 (ANO5) protein expression is absent in patients with the founder mutation (c.191dupA, p.Asn64Lysfs*15) (2, 6, 7) and ANO5 protein is reduced in patients with point mutations (e.g. c.2101A>G, p.N701D and c.2272C>T, p.R758C) (7,8). Whereas ANO5-myopathies are inherited in a recessive manner, dominant (probably gain of function) ANO5 mutations are associated with gnathodiaphyseal dysplasia, a skeletal syndrome characterized by cementosseous lesions of the jaw, osteomyelitis, bone fragility (9, 10) and dental tumors (11-13).ANO5 is a member of the TMEM16/Anoctamin family that includes ion channels and phospholipid scramblases (14-16).…”

mentioning

confidence: 99%

“…Furthermore, phospholipid scrambling is absent in muscle precursor cells from ANO5-KO mice despite expression of ANO6 (21), suggesting a unique, muscle-specific role for ANO5mediated phospholipid scrambling. However, the pathologic mechanism of ANO5 myopathies is poorly understood and it remains unclear whether ANO5-linked disorders are a consequence of defective PLS.One suggestion is that ANO5 myopathies are caused by defects in the ability of muscle fibers to self-repair (1,8,24,25). Skeletal muscle endures considerable mechanical stress even during normal use, which can produce small tears in the plasma membrane (sarcolemma) (26).…”

mentioning

confidence: 99%

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Defective Trafficking of Annexins to the Site of Injury in ANO5-Knockout Muscle Fibers

Foltz

Cui

Choo

et al. 2020

Preprint

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Mutations in ANO5 (TMEM16E) cause limb-girdle muscular dystrophy R12 (limb-girdle muscular dystrophy type 2L). Recent evidence implicates defective plasma membrane repair as a likely mechanism for the disorder. Here, we probe the ANO5-dependency of the membrane repair pathway using a laser wounding assay in Ano5 knockout mouse muscle fibers. Wounded myofibers from Ano5 knockout mice exhibit delayed membrane resealing relative to wild type fibers as revealed by an increased uptake of the membrane-impermeant FM1-43 dye and a prolonged elevation of intracellular Ca 2+ . The trafficking of several annexin proteins, which together form a cap at the site of injury, is altered in Ano5 knockout fibers. Annexin A2 accumulates at the wound to nearly twice the level observed in WT fibers, while annexin A6 accumulation is substantially inhibited in the absence of ANO5. Furthermore, trafficking of annexins A1 and A5 to the cap is decreased in the Ano5 knockout. These changes are correlated with an alteration in the fine structure of the annexin repair cap and the shedding of annexin-positive extracellular vesicles. Our results suggest that the meticulous coordination of the annexin repair machinery required to effectively reseal wounded sarcolemma is disrupted in Ano5 knockout mice. ANO5 is a putative phospholipid scramblase, responsible for exposure of intracellular phospholipids to the extracellular leaflet of the plasma membrane. However, because the membrane repair defect is rescued by overexpression of wild type ANO5 or a scramblase-defective mutant, we suggest that ANO5-mediated phospholipid scrambling is not essential for membrane repair. Significance StatementMutations in ANO5/TMEM16E cause myopathies of variable severity, with some patients losing ambulation entirely. Unfortunately, relatively little is known about the function of ANO5 at the protein level, but it has been suggested that ANO5 plays a role in the repair of injured muscle plasma membranes. Here, we investigate the mechanism of ANO5-mediated repair and find that annexin proteins, which in normal muscle form a cap to seal wounds, traffic abnormally to the cap when ANO5 is not expressed. Muscle fibers lacking ANO5 reseal more slowly and thus are exposed to prolonged intracellular calcium elevation that can damage the fibers. Our findings contribute to the growing literature implicating failed repair as a probable pathogenic mechanism in patients with ANO5 mutations.Recessive mutations in the ANO5 gene are responsible for a spectrum of myopathies with variable severity. These disorders, which are characterized by muscle weakness and atrophy, include limb girdle muscular dystrophy R12 (LGMDR12/LGMD2L), Miyoshi muscular dystrophy type 3 (MMD3), and muscle weakness with myalgia and rhabdomyolysis (1-4). Since ANO5-myopathies were discovered in 2010 (1), >70 ANO5 variants have been reported to be pathogenic (https://www.ncbi.nlm.nih.gov/clinvar/). In a recent screen of 35 LGMD genes in 4656 clinically-suspected LGMD patients, ANO5 was the 4 th most likely cont...

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Section: Discussionsupporting

confidence: 93%

Section: Introductionmentioning

confidence: 77%