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Fig. 3
Normal laminin deposition/organization is necessary for concentration of Mmp11 at MTJs. A1–A3 Anterior left, dorsal top, side-mounted, wild-type embryos stained with anti-Mmp11 antibody at 20, 26, or 48 hpf, respectively. Note that Mmp11 increasingly concentrates at MTJs (white arrows) over developmental time. B1, B2 Western blots of whole embryo homogenates at 26 hpf using an anti-Mmp11 antibody. B1 Laminin gamma1 morphants have approximately 8.5-fold less Mmp11 protein compared to controls. B2 Laminin gamma1 mutants show no change (approximately 1.2-fold more) Mmp11 protein compared to siblings. (C–E) Side view, anterior left, dorsal top, 27 hpf embryos stained with phalloidin (light blue), and anti-Mmp11 antibody (purple). C1–C3 Control embryo. Mmp11 localizes to the MTJ (peaks in C3) and is absent from the myotome (valleys in c3). c3 Histogram of relative pixel intensities across the white rectangle in panel c2. d1–d3 nrk2b morphant. Mmp11 is expressed and concentrates at MTJ in nrk2b morphants (white arrow in d2 and peaks in d3); however, Mmp11 localization is disrupted as can be seen by the increased staining within myotomes (higher intensity of valleys in d3 compared to c3) (n = 13 embryos). d3 Histogram of relative pixel intensities across the white rectangle in panel D2. E1–E3 laminin gamma1 mutant. Mmp11 does not localize to MTJs (absence of clear peaks and valleys in E3), except in small patches (white arrowhead in E2) (n = 7 embryos). E3 Histogram of relative pixel intensities across the white rectangle in panel E2. Scale bars are 50 μm