Figure 1.

Targeting properties of the two-domain Ca²⁺ channel fragments GFP-I⋅II and III⋅IV expressed in dysgenic myotubes. Transfected myotubes were double-immunofluorescence labeled with anti-GFP to detect GFP-I⋅II, anti-α_1S to detect III⋅IV (the locations of epitopes are indicated in green in the schematic drawings of α_1S fragments below the micrographs), and with anti-RyR as independent triad marker. When coexpressed, GFP-I⋅II and III⋅IV are colocalized with one another (first column) and with the RyR1 (second column) in clusters corresponding to triad junctions (examples indicated with arrows). Individually expressed GFP-I⋅II (third column) and III⋅IV (fourth column) are not colocalized with RyR1 clusters but are expressed throughout the endoplasmic reticulum (ER)/SR system. Merged color images (bottom row) of the micrographs above show colocalization of red and green fluorescent antibodies as yellow foci and lack of colocalization as separate red and green structures. The schematic drawings show the repetitive transmembrane domain structure of the α_1S fragments expressed in the myotubes shown above. N, nuclei. (Scale bar, 20 μm.)