Abstract
Sarcospan (SSPN) is a core component of the dystrophin–glycoprotein complex (DGC). Multiple SSPN transcripts are ubiquitously expressed and SSPN splicing is disrupted in many lung tumors, suggesting the importance of SSPN-related mRNAs. We describe the isolation of an alternatively spliced isoform of SSPN, which we designate ‘microspan’ based on its small size relative to SSPN. Microspan has two transmembrane domains and a novel C-terminus. We demonstrate that microspan is not an integral component of the DGC and is not perturbed by the loss of dystrophin. Microspan protein is detected at the sarcoplasmic reticulum (SR) using indirect immunofluorescence and immunoelectron microscopy. Furthermore, microspan purifies with skeletal muscle SR membranes and not transverse tubules. Mice engineered to over-express microspan display severe kyphosis and die at approximately 8 weeks of age. Levels of ryanodine receptor, dihydropyridine receptor, and SERCA-1 are greatly reduced in microspan transgenic muscle. Furthermore, electron microscopy reveals that microspan over-expression causes a dramatic perturbation in triad structure. Our findings suggest that microspan is an important component of the SR and may contribute to excitation–contraction coupling.
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Abbreviations
- AR-LGMD:
-
autosomal recessive limb girdle muscular dystrophy
- CRUs:
-
calcium release units
- DGC:
-
dystrophin–glycoprotein complex
- DGs:
-
dystroglycans
- DHPR:
-
α1-dihydropyridine receptor
- DMD:
-
Duchenne muscular dystrophy
- EDL:
-
extensor digitorum longus
- LEL:
-
large extracellular loop
- PVDF:
-
polyvinylidene fluoride
- RyR:
-
ryanodine receptor
- SEL:
-
small extracellular loop
- SGs:
-
sarcoglycans
- SR:
-
sarcoplasmic reticulum
- SSPN:
-
sarcospan
- TA:
-
tibalis anterior
- Tg:
-
transgenic
- TM:
-
transmembrane domain
- T-tubules:
-
transverse tubules
- μSPN:
-
microspan
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Acknowledgements
We thank Dr. Kevin P. Campbell (HHMI, University of Iowa College of Medicine) for DHPR and RyR antibodies, Dr. Jeffrey S. Chamberlain (University of Washington, Seattle) for transgenic expression plasmids, Birgitta Sjostrand, Marianne Cilluffo and Suni Allen (Brain Research Institute electron microscopy core, UCLA) for EM experiments and Matt Schibler (Carol Moss Spivak Cell Imaging Facility, UCLA) for confocal microscopy. We are indebted to Dr. Melissa J. Spencer (UCLA) for guidance with transgene construction. A.K. Peter was supported by the Molecular, Cellular, and Integrative Physiology predoctoral training fellowship (NIH:T32GM65823). This work was supported by the Muscular Dystrophy Association (MDA3704 to G.M.), the Roy Castle Lung Cancer Foundation, UK (J.H.) and the NIH (AR48179-01 to R.H.C.).
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Miller, G., Peter, A.K., Espinoza, E. et al. Over-expression of Microspan, a novel component of the sarcoplasmic reticulum, causes severe muscle pathology with triad abnormalities. J Muscle Res Cell Motil 27, 545–558 (2006). https://doi.org/10.1007/s10974-006-9069-2
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DOI: https://doi.org/10.1007/s10974-006-9069-2