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Fig. 4 pMRLC is abnormally elevated and persists apically in the developing brain of mypt1 mutants. (A) Mypt1 as part of myosin phosphatase. (B) Hypothesis for effect of mypt1 mutant myosin phosphatase. (C) Western blot for pMRLC brain tissue dissected from zebrafish embryos at 18, 21 and 24 hpf after injection with control MO or mypt1 MO, with actin control. Approximately 100 brains were dissected for each treatment group and 50 μg of protein was analyzed. (D) Quantitation of fold changes of pMRLC normalized to actin at 18 hpf, comparing control MO-injected with mypt1 MO-injected embryos. Results are representative of three independent experiments. (E) Quantitation of fold changes of pMRLC normalized to actin over time in control MO embryos. Results are representative of three independent experiments. (F-Q) Transverse vibratome sections through the hindbrain of embryos stained for pMRLC (green) and counterstained for nuclei (propidium iodide, red). Representative images from four independent experiments; two to three embryos were imaged at each stage per experiment. Wild-type embryo at 19 (F,G), 21 (H,I) and 24 (J,K) hpf. mypt1 mutant embryo at 19 (L,M), 21 (N,O) and 24 (P,Q) hpf. (G,I,K,M,O,Q) Higher magnification of the boxed region in F,H,J,L,N,P. Dorsal is up. Arrows point to the apical neuroepithelial surface in all images. Scale bars: 50 μm in F,H,J,L,N,P; 10 μm in G,I,K,M,O,Q.