Fig. 2

Two isoforms of p120 catenin proteins are detected in homogenates of embryos between 3.7 and 38 hpf in SDS-PAGE Western blots using a polyclonal antibody to p120 catenin. SDS-PAGE homogenates from 3 embryos were added to each lane. Stages shown are: oblong about 4,000 cells (3.7 hpf); shield (6 hpf); bud at the end of epiboly (11 hpf); 2-somite stage (11 hpf); 10-somite stage (14 hpf); 21-somite stage (19.5 hpf); Prim 5 (24 hpf); Prim 25 (38 hpf). The p120 catenin protein that is initiated at Met-1 has a Mr of 120 k, and the protein initiated at Met-3 has a Mr of 100 k. B: Injection of antisense splice morpholino oligonucleotides (Sp-MO) blocks new mRNA synthesis and decreases synthesis of 100-kDa p120 catenin proteins initiating at Met-3. Zebrafish embryos were collected at the 1-cell stage and injected with 6 or 8 ng of δ 1 splice-MO. Zebrafish embryos were counted, and quantitatively lysed in SDS-PAGE buffer as described in the Experimental Procedures section resulting in the loading of 1-embryo per lane. Uninjected embryos were homogenized at shield (6 hpf), 80 to 90% epiboly (9 hpf), and bud (10 hpf) and δ1 splice-MO-injected zebrafish embryos were homogenized at shield and bud. All samples were analyzed by SDS-PAGE Western blot with a polyclonal anti-p120 catenin antibody. C: Abundance of p120 was measured as described in the Experimental Procedures section and plotted on a bar graph. The Y-axis represents relative density in arbitrary units determined by ImageJ with a background correction using Rolling Ball Subtraction.