Fig. 4. Subcellular localization of mutant plectin fragments containing a β-synemin-binding site in transfected C2C12 myoblasts and myotubes. C2C12 myoblasts were transfected with plasmids expressing Myc-tagged plectin Ex 1, CHD and PlD-M fragments, and then cultured in differentiation medium. At 60-84 hours post transfection, differentiated myotubes were immunolabeled with antibodies specific for Myc and β-synemin. Localization of β-synemin along SFLSs and throughout the sarcoplasm in a dotted pattern was found in control myotubes doubly immunolabeled with Alxea-Fluor-594-conjugated phallotoxins and anti-β-synemin pAb (A-F). The three mutant plectin fragments more or less colocalized with β-synemin along SFLS (arrows in J-L,P-R,V-X). Arrowheads in V and X indicate sites associated predominantly with β-synemin, but scarcely with PlD-M. Bars, 10 µm.