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Fig. 7. 
-DG is destabilized in phenotypic SSPN-Tg muscle. (A) Schematic diagram showing the steps in the SDS titration experiment. Skeletal muscle from non-Tg and SSPN-Tg mice was subjected to sequential extractions in lysis buffer with increasing SDS concentrations. Skeletal muscle was homogenized in 0.25% SDS and centrifuged to separate soluble (supe) from insoluble (pellet) proteins. The pellet was resuspended in a 0.5% SDS buffer for a second round of protein extraction. This process was repeated with 1%, 2% and 3% SDS. (B) Protein supernatants from each SDS titration were separated using SDS-polyacrylamide gels and transferred to nitrocellulose membranes. Immunoblots were stained with antibodies to laminin (Lam), - and 
-DG, -, -, and 
-SG, hSSPN, and mSSPN as indicated. 1% SDS was required to extract -DG in non-Tg muscle, which represents a strong and stable attachment of -DG to the DGC. In non-phenotypic SSPN-Tg muscle, -DG is removed under lower stringency conditions (0.5% SDS), suggesting that -DG is weakened by presence of SSPN overexpression. Finally, -DG is readily extracted from phenotypic SSPN-Tg muscle with 0.25% SDS, supporting the hypothesis that SSPN destabilizes -DG. (C) Densitometry of -DG staining from immunoblots. Relative levels of -DG found in the supernatant fraction after treatment with 0.25%, 0.50% and 1.0% was quantified for all muscle samples analyzed. (D) Schematic models illustrating -DG attachment to the DGC in non-Tg, non-phenotypic SSPN-Tg, and phenotypic SSPN-Tg.
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