|
Fig. 5 Itga6 functions in the Nrk2b-NAD+-laminin pathway and is required for NAD+-mediated rescue of MTJ morphogenesis and dystrophy.
(A-C, E-F, H-I) Anterior left, dorsal top, side-mounted, 2 dpf embryos stained with phalloidin (white) to visualize actin. In pseudocolored panels (A1, B1, C, E, F), MTJ boundaries are blue, and abnormally long muscle fibers are red. (A) MTJs are V-shaped and continuous in control embryos. (B) In itga6 morphants, MTJs are U-shaped, discontinuous, and crossed by abnormally long muscle fibers (red arrows). (C) Co-injection of p53 MOs does not rescue MTJ failure in itga6 morphants. (D) Quantification of MTJ failure at 2 dpf in controls, itga6 morphants, and itga6;p53 double morphants. (E–F) Co-injection of itga6 cDNA that does not contain the MO target sites with itga6 MOs rescues the itga6 morphant phenotype. (G) Quantification of MTJ failure shows that NAD+ treatment does not rescue MTJ failure in itga6 morphants, suggesting that NAD+ requires Itga6 for rescue of MTJ failure. (H) dag1;itga6 double morphants have U-shaped MTJs and dystrophy (white arrowheads). (I) NAD+ does not reduce MTJ angles (not shown) or dystrophy in dag1;itga6 double morphants, suggesting that Itga6 is also required for NAD+-mediated rescue of MTJ angles and dystrophy. (J) Quantification of dystrophy shows significant rescue by exogenous NAD+ in dag1 morphants, but not dag1;itga6 double morphants; *p<0.05; N.S., not significant. Scale bars are 50 micrometers.