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Zebrafish models for nemaline myopathy reveal a spectrum of nemaline bodies contributing to reduced muscle function.

Sztal TE, Zhao M, Williams C, Oorschot V, Parslow AC, Giousoh A, Yuen M, Hall TE, Costin A, Ramm G, Bird PI, Busch-Nentwich EM, Stemple DL, Currie PD, Cooper ST, Laing NG, Nowak KJ, Bryson-Richardson RJ - Acta Neuropathol. (2015)

Bottom Line: Another subtype results from a reduction of actin and forms a more stable cytoplasmic body.In contrast, the final type originates at the Z-disk and is associated with myofibrillar disorganization.In addition, we show that the ACTA1(D286G) mutation causes impaired actin incorporation and localization in the sarcomere.

View Article: PubMed Central - PubMed

Affiliation: School of Biological Sciences, Monash University, Melbourne, VIC, Australia.

ABSTRACT
Nemaline myopathy is characterized by muscle weakness and the presence of rod-like (nemaline) bodies. The genetic etiology of nemaline myopathy is becoming increasingly understood with mutations in ten genes now known to cause the disease. Despite this, the mechanism by which skeletal muscle weakness occurs remains elusive, with previous studies showing no correlation between the frequency of nemaline bodies and disease severity. To investigate the formation of nemaline bodies and their role in pathogenesis, we generated overexpression and loss-of-function zebrafish models for skeletal muscle α-actin (ACTA1) and nebulin (NEB). We identify three distinct types of nemaline bodies and visualize their formation in vivo, demonstrating these nemaline bodies not only exhibit different subcellular origins, but also have distinct pathological consequences within the skeletal muscle. One subtype is highly dynamic and upon breakdown leads to the accumulation of cytoplasmic actin contributing to muscle weakness. Examination of a Neb-deficient model suggests this mechanism may be common in nemaline myopathy. Another subtype results from a reduction of actin and forms a more stable cytoplasmic body. In contrast, the final type originates at the Z-disk and is associated with myofibrillar disorganization. Analysis of zebrafish and muscle biopsies from ACTA1 nemaline myopathy patients demonstrates that nemaline bodies also possess a different protein signature. In addition, we show that the ACTA1(D286G) mutation causes impaired actin incorporation and localization in the sarcomere. Together these data provide a novel examination of nemaline body origins and dynamics in vivo and identifies pathological changes that correlate with muscle weakness.

No MeSH data available.


Related in: MedlinePlus

a) qRT-PCR analysis of skeletal α-actin genes in zebrafish tail muscle at 2 dpf. Error bars represent ±SEM for three replicate experiments with each experiment comprising a pooled samples of 20 fish, *p < 0.01. b, c The decrease in total amount of α-actin protein in Actc1b (ex 2 and UTR) morphants at 2 dpf was confirmed and quantitated by Western blot. β-tubulin was used as a loading control
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Fig9: a) qRT-PCR analysis of skeletal α-actin genes in zebrafish tail muscle at 2 dpf. Error bars represent ±SEM for three replicate experiments with each experiment comprising a pooled samples of 20 fish, *p < 0.01. b, c The decrease in total amount of α-actin protein in Actc1b (ex 2 and UTR) morphants at 2 dpf was confirmed and quantitated by Western blot. β-tubulin was used as a loading control

Mentions: Using our zebrafish system, we also created a model for recessive nemaline myopathy. Zebrafish possess two skeletal muscle α-actin genes (acta1a and acta1b) and two cardiac α-actin genes (actc1a and actc1b), which are all expressed during early muscle development. qRT-PCR analyses showed that actc1b is the predominant isoform expressed in the skeletal muscle at 2 dpf (Fig. 9a). Thus, we chose to knockdown Actc1b to reduce the amount of α-actin in skeletal muscle.Fig. 9


Zebrafish models for nemaline myopathy reveal a spectrum of nemaline bodies contributing to reduced muscle function.

Sztal TE, Zhao M, Williams C, Oorschot V, Parslow AC, Giousoh A, Yuen M, Hall TE, Costin A, Ramm G, Bird PI, Busch-Nentwich EM, Stemple DL, Currie PD, Cooper ST, Laing NG, Nowak KJ, Bryson-Richardson RJ - Acta Neuropathol. (2015)

a) qRT-PCR analysis of skeletal α-actin genes in zebrafish tail muscle at 2 dpf. Error bars represent ±SEM for three replicate experiments with each experiment comprising a pooled samples of 20 fish, *p < 0.01. b, c The decrease in total amount of α-actin protein in Actc1b (ex 2 and UTR) morphants at 2 dpf was confirmed and quantitated by Western blot. β-tubulin was used as a loading control
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4541704&req=5

Fig9: a) qRT-PCR analysis of skeletal α-actin genes in zebrafish tail muscle at 2 dpf. Error bars represent ±SEM for three replicate experiments with each experiment comprising a pooled samples of 20 fish, *p < 0.01. b, c The decrease in total amount of α-actin protein in Actc1b (ex 2 and UTR) morphants at 2 dpf was confirmed and quantitated by Western blot. β-tubulin was used as a loading control
Mentions: Using our zebrafish system, we also created a model for recessive nemaline myopathy. Zebrafish possess two skeletal muscle α-actin genes (acta1a and acta1b) and two cardiac α-actin genes (actc1a and actc1b), which are all expressed during early muscle development. qRT-PCR analyses showed that actc1b is the predominant isoform expressed in the skeletal muscle at 2 dpf (Fig. 9a). Thus, we chose to knockdown Actc1b to reduce the amount of α-actin in skeletal muscle.Fig. 9

Bottom Line: Another subtype results from a reduction of actin and forms a more stable cytoplasmic body.In contrast, the final type originates at the Z-disk and is associated with myofibrillar disorganization.In addition, we show that the ACTA1(D286G) mutation causes impaired actin incorporation and localization in the sarcomere.

View Article: PubMed Central - PubMed

Affiliation: School of Biological Sciences, Monash University, Melbourne, VIC, Australia.

ABSTRACT
Nemaline myopathy is characterized by muscle weakness and the presence of rod-like (nemaline) bodies. The genetic etiology of nemaline myopathy is becoming increasingly understood with mutations in ten genes now known to cause the disease. Despite this, the mechanism by which skeletal muscle weakness occurs remains elusive, with previous studies showing no correlation between the frequency of nemaline bodies and disease severity. To investigate the formation of nemaline bodies and their role in pathogenesis, we generated overexpression and loss-of-function zebrafish models for skeletal muscle α-actin (ACTA1) and nebulin (NEB). We identify three distinct types of nemaline bodies and visualize their formation in vivo, demonstrating these nemaline bodies not only exhibit different subcellular origins, but also have distinct pathological consequences within the skeletal muscle. One subtype is highly dynamic and upon breakdown leads to the accumulation of cytoplasmic actin contributing to muscle weakness. Examination of a Neb-deficient model suggests this mechanism may be common in nemaline myopathy. Another subtype results from a reduction of actin and forms a more stable cytoplasmic body. In contrast, the final type originates at the Z-disk and is associated with myofibrillar disorganization. Analysis of zebrafish and muscle biopsies from ACTA1 nemaline myopathy patients demonstrates that nemaline bodies also possess a different protein signature. In addition, we show that the ACTA1(D286G) mutation causes impaired actin incorporation and localization in the sarcomere. Together these data provide a novel examination of nemaline body origins and dynamics in vivo and identifies pathological changes that correlate with muscle weakness.

No MeSH data available.


Related in: MedlinePlus