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Loss of myotubularin function results in T-tubule disorganization in zebrafish and human myotubular myopathy.

Dowling JJ, Vreede AP, Low SE, Gibbs EM, Kuwada JY, Bonnemann CG, Feldman EL - PLoS Genet. (2009)

Bottom Line: Zebrafish with reduced levels of myotubularin have significantly impaired motor function and obvious histopathologic changes in their muscle.We demonstrate for the first time that myotubularin functions to regulate PI3P levels in a vertebrate in vivo, and that homologous myotubularin-related proteins can functionally compensate for the loss of myotubularin.Based on our findings, we speculate that congenital myopathies, usually considered entities with similar clinical features but very disparate pathomechanisms, may at their root be disorders of calcium homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of Michigan Medical Center, Ann Arbor, Michigan, USA. jamedowl@umich.edu

ABSTRACT
Myotubularin is a lipid phosphatase implicated in endosomal trafficking in vitro, but with an unknown function in vivo. Mutations in myotubularin cause myotubular myopathy, a devastating congenital myopathy with unclear pathogenesis and no current therapies. Myotubular myopathy was the first described of a growing list of conditions caused by mutations in proteins implicated in membrane trafficking. To advance the understanding of myotubularin function and disease pathogenesis, we have created a zebrafish model of myotubular myopathy using morpholino antisense technology. Zebrafish with reduced levels of myotubularin have significantly impaired motor function and obvious histopathologic changes in their muscle. These changes include abnormally shaped and positioned nuclei and myofiber hypotrophy. These findings are consistent with those observed in the human disease. We demonstrate for the first time that myotubularin functions to regulate PI3P levels in a vertebrate in vivo, and that homologous myotubularin-related proteins can functionally compensate for the loss of myotubularin. Finally, we identify abnormalities in the tubulo-reticular network in muscle from myotubularin zebrafish morphants and correlate these changes with abnormalities in T-tubule organization in biopsies from patients with myotubular myopathy. In all, we have generated a new model of myotubular myopathy and employed this model to uncover a novel function for myotubularin and a new pathomechanism for the human disease that may explain the weakness associated with the condition (defective excitation-contraction coupling). In addition, our findings of tubuloreticular abnormalities and defective excitation-contraction coupling mechanistically link myotubular myopathy with several other inherited muscle diseases, most notably those due to ryanodine receptor mutations. Based on our findings, we speculate that congenital myopathies, usually considered entities with similar clinical features but very disparate pathomechanisms, may at their root be disorders of calcium homeostasis.

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Related in: MedlinePlus

Abnormal motor function in myotubularin morphants.(A) Quantitation of spontaneous embryo coiling at 24 hpf (see also Supplemental Videos 1 and 2). On average, CTL morphants coiled 10.2 times in 15 seconds, while MTM morphants coiled only 5.2 times. (B) Quantitation of chorion hatching in 60 hpf morphants. 87.2% of CTL morphants are hatched from their protective chorions by 60 hpf, as opposed to only 35.3% of MTM morphants. (C) Touch-evoked swimming was video captured in 72 hpf morphants. As expected, CTL morphants responded to tactile stimuli with a rapid escape response contraction followed by swimming. Conversely, MTM morphants displayed a weak escape contraction followed by “twittering” movements (example at 33 ms) but never normal swimming. Scale bar = 1 mm.
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pgen-1000372-g002: Abnormal motor function in myotubularin morphants.(A) Quantitation of spontaneous embryo coiling at 24 hpf (see also Supplemental Videos 1 and 2). On average, CTL morphants coiled 10.2 times in 15 seconds, while MTM morphants coiled only 5.2 times. (B) Quantitation of chorion hatching in 60 hpf morphants. 87.2% of CTL morphants are hatched from their protective chorions by 60 hpf, as opposed to only 35.3% of MTM morphants. (C) Touch-evoked swimming was video captured in 72 hpf morphants. As expected, CTL morphants responded to tactile stimuli with a rapid escape response contraction followed by swimming. Conversely, MTM morphants displayed a weak escape contraction followed by “twittering” movements (example at 33 ms) but never normal swimming. Scale bar = 1 mm.

Mentions: In zebrafish, the first recognizable muscle dependent motor function, detected between 17 and 26 hpf, is spontaneous embryo coiling [21]. On average, control injected embryos had 10.2 (+/−0.4) spontaneous muscle contractions per 15 second period (Supplemental Video 1). Conversely, embryos injected with myotubularin morpholinos had only 5.2 (+/−0.5) contractions in the same period (Figure 2A and Supplemental Video 2). This abnormality was highly reproducible (P<0.0001), and marked the earliest observed functional abnormality in zebrafish with reduced myotubularin levels.


Loss of myotubularin function results in T-tubule disorganization in zebrafish and human myotubular myopathy.

Dowling JJ, Vreede AP, Low SE, Gibbs EM, Kuwada JY, Bonnemann CG, Feldman EL - PLoS Genet. (2009)

Abnormal motor function in myotubularin morphants.(A) Quantitation of spontaneous embryo coiling at 24 hpf (see also Supplemental Videos 1 and 2). On average, CTL morphants coiled 10.2 times in 15 seconds, while MTM morphants coiled only 5.2 times. (B) Quantitation of chorion hatching in 60 hpf morphants. 87.2% of CTL morphants are hatched from their protective chorions by 60 hpf, as opposed to only 35.3% of MTM morphants. (C) Touch-evoked swimming was video captured in 72 hpf morphants. As expected, CTL morphants responded to tactile stimuli with a rapid escape response contraction followed by swimming. Conversely, MTM morphants displayed a weak escape contraction followed by “twittering” movements (example at 33 ms) but never normal swimming. Scale bar = 1 mm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2631153&req=5

pgen-1000372-g002: Abnormal motor function in myotubularin morphants.(A) Quantitation of spontaneous embryo coiling at 24 hpf (see also Supplemental Videos 1 and 2). On average, CTL morphants coiled 10.2 times in 15 seconds, while MTM morphants coiled only 5.2 times. (B) Quantitation of chorion hatching in 60 hpf morphants. 87.2% of CTL morphants are hatched from their protective chorions by 60 hpf, as opposed to only 35.3% of MTM morphants. (C) Touch-evoked swimming was video captured in 72 hpf morphants. As expected, CTL morphants responded to tactile stimuli with a rapid escape response contraction followed by swimming. Conversely, MTM morphants displayed a weak escape contraction followed by “twittering” movements (example at 33 ms) but never normal swimming. Scale bar = 1 mm.
Mentions: In zebrafish, the first recognizable muscle dependent motor function, detected between 17 and 26 hpf, is spontaneous embryo coiling [21]. On average, control injected embryos had 10.2 (+/−0.4) spontaneous muscle contractions per 15 second period (Supplemental Video 1). Conversely, embryos injected with myotubularin morpholinos had only 5.2 (+/−0.5) contractions in the same period (Figure 2A and Supplemental Video 2). This abnormality was highly reproducible (P<0.0001), and marked the earliest observed functional abnormality in zebrafish with reduced myotubularin levels.

Bottom Line: Zebrafish with reduced levels of myotubularin have significantly impaired motor function and obvious histopathologic changes in their muscle.We demonstrate for the first time that myotubularin functions to regulate PI3P levels in a vertebrate in vivo, and that homologous myotubularin-related proteins can functionally compensate for the loss of myotubularin.Based on our findings, we speculate that congenital myopathies, usually considered entities with similar clinical features but very disparate pathomechanisms, may at their root be disorders of calcium homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of Michigan Medical Center, Ann Arbor, Michigan, USA. jamedowl@umich.edu

ABSTRACT
Myotubularin is a lipid phosphatase implicated in endosomal trafficking in vitro, but with an unknown function in vivo. Mutations in myotubularin cause myotubular myopathy, a devastating congenital myopathy with unclear pathogenesis and no current therapies. Myotubular myopathy was the first described of a growing list of conditions caused by mutations in proteins implicated in membrane trafficking. To advance the understanding of myotubularin function and disease pathogenesis, we have created a zebrafish model of myotubular myopathy using morpholino antisense technology. Zebrafish with reduced levels of myotubularin have significantly impaired motor function and obvious histopathologic changes in their muscle. These changes include abnormally shaped and positioned nuclei and myofiber hypotrophy. These findings are consistent with those observed in the human disease. We demonstrate for the first time that myotubularin functions to regulate PI3P levels in a vertebrate in vivo, and that homologous myotubularin-related proteins can functionally compensate for the loss of myotubularin. Finally, we identify abnormalities in the tubulo-reticular network in muscle from myotubularin zebrafish morphants and correlate these changes with abnormalities in T-tubule organization in biopsies from patients with myotubular myopathy. In all, we have generated a new model of myotubular myopathy and employed this model to uncover a novel function for myotubularin and a new pathomechanism for the human disease that may explain the weakness associated with the condition (defective excitation-contraction coupling). In addition, our findings of tubuloreticular abnormalities and defective excitation-contraction coupling mechanistically link myotubular myopathy with several other inherited muscle diseases, most notably those due to ryanodine receptor mutations. Based on our findings, we speculate that congenital myopathies, usually considered entities with similar clinical features but very disparate pathomechanisms, may at their root be disorders of calcium homeostasis.

Show MeSH
Related in: MedlinePlus