Fig. 7: Location of Nogo-A in mature skeletal muscle and C2C12 myoblasts, and identification of Nogo-A interacting factors during differentiation.

A–C Localization of Nogo-A in mature skeletal muscle and C2C12 myoblasts. A Isolated gastrocnemius muscles from 8-week-old C27BL/6 mouse were sectioned and stained for Nogo-A (green) and MYH2 (red). Scale bar = 100 µm. B IF staining of Nogo-A (red) and calnexin (ER protein, green) in growth-conditioned C2C12 cells. Scale bar = 10 µm. C IF staining of Nogo-A (red) and desmin (muscle-specific, type III intermediate filament, green) in differentiation-conditioned C2C12 cells. Scale bar = 10 µm. D–F Expression levels of Nogo-A-interacting molecules suggested by immunoprecipitation-mass spectrometry analysis of differentiated C2C12. qRT-PCR analysis of indicated genes in C2C12 cells cultured with DM for 2 or 4 days (D) and in muscles after 3 days or 2 weeks of notexin injury (E). F Immunoprecipitation with a Nogo-A-specific antibody was performed using lysates of C2C12 cells maintained in GM or DM for 4 days. WB analysis of indicated proteins in immunoprecipitated samples. Molecular weights (kDa) are indicated. G Quantitative assessment of band intensities of blots in F using the NIH ImageJ software. H Filamin-mediated cell motility regulation and proposed model of Nogo-A’s role in muscle differentiation via its interaction with filamin-C.