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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

T-tubule structural abnormalities in myotubularin morphant muscle.T-tubule (vertical arrows) and sarcoplasmic reticulum (angled arrows) abnormalities as demonstrated by electron microscopy. Control morphant (panel 1; CTL): normal T-tubule triad with accompanying thin, well-organized SR network (arrow). Myotubularin morphants (panels 2–4): Panel 2 shows mildly dilated triads and SR networks. Panel 3 shows severely dilated and dysmorphic triads and widely looped SR. Panel 4 illustrates severe disorganization, with unrecognizable T-tubule triads and aberrant adjacent SR networks. Scale bar = 500 nm.
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pgen-1000372-g009: T-tubule structural abnormalities in myotubularin morphant muscle.T-tubule (vertical arrows) and sarcoplasmic reticulum (angled arrows) abnormalities as demonstrated by electron microscopy. Control morphant (panel 1; CTL): normal T-tubule triad with accompanying thin, well-organized SR network (arrow). Myotubularin morphants (panels 2–4): Panel 2 shows mildly dilated triads and SR networks. Panel 3 shows severely dilated and dysmorphic triads and widely looped SR. Panel 4 illustrates severe disorganization, with unrecognizable T-tubule triads and aberrant adjacent SR networks. Scale bar = 500 nm.

Mentions: Based on this localization, we were interested in the effect of myotubularin knockdown on T-tubule organization. We performed ultrastructural analysis using electron microscopy (Figure 9). Muscle from control morpholino injected embryos exhibited the normal pattern of T-tubules and sarcoplasmic reticulum (SR), with the SR coursing tightly through the sarcomeres and the T-tubules forming triads at regular periods. Conversely, muscle from myotubularin morpholino injected embryos had grossly aberrant SR and T-tubule networks (Figure 9). The SR networks were irregular, disorganized, and often randomly interspersed throughout the sarcomere. The T-tubule triads showed a range of abnormalities, from mild changes in electron density of the triad (arrow, upper right panel), to severe dilation of the triad structure (arrows, lower right panel), to fibers with essentially unrecognizable SR/triad areas (arrow, lower left panel).


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)

T-tubule structural abnormalities in myotubularin morphant muscle.T-tubule (vertical arrows) and sarcoplasmic reticulum (angled arrows) abnormalities as demonstrated by electron microscopy. Control morphant (panel 1; CTL): normal T-tubule triad with accompanying thin, well-organized SR network (arrow). Myotubularin morphants (panels 2–4): Panel 2 shows mildly dilated triads and SR networks. Panel 3 shows severely dilated and dysmorphic triads and widely looped SR. Panel 4 illustrates severe disorganization, with unrecognizable T-tubule triads and aberrant adjacent SR networks. Scale bar = 500 nm.
© Copyright Policy
Related In: Results  -  Collection

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

pgen-1000372-g009: T-tubule structural abnormalities in myotubularin morphant muscle.T-tubule (vertical arrows) and sarcoplasmic reticulum (angled arrows) abnormalities as demonstrated by electron microscopy. Control morphant (panel 1; CTL): normal T-tubule triad with accompanying thin, well-organized SR network (arrow). Myotubularin morphants (panels 2–4): Panel 2 shows mildly dilated triads and SR networks. Panel 3 shows severely dilated and dysmorphic triads and widely looped SR. Panel 4 illustrates severe disorganization, with unrecognizable T-tubule triads and aberrant adjacent SR networks. Scale bar = 500 nm.
Mentions: Based on this localization, we were interested in the effect of myotubularin knockdown on T-tubule organization. We performed ultrastructural analysis using electron microscopy (Figure 9). Muscle from control morpholino injected embryos exhibited the normal pattern of T-tubules and sarcoplasmic reticulum (SR), with the SR coursing tightly through the sarcomeres and the T-tubules forming triads at regular periods. Conversely, muscle from myotubularin morpholino injected embryos had grossly aberrant SR and T-tubule networks (Figure 9). The SR networks were irregular, disorganized, and often randomly interspersed throughout the sarcomere. The T-tubule triads showed a range of abnormalities, from mild changes in electron density of the triad (arrow, upper right panel), to severe dilation of the triad structure (arrows, lower right panel), to fibers with essentially unrecognizable SR/triad areas (arrow, lower left panel).

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