Figure 9

Two-dimensional SDS-PAGE of embryonic heart proteins followed by Western Blotting using CH1 (anti-tropomyosin) antibody. Untreated stage 36 normal hearts (A); Untreated stage 36 mutant hearts (B); Enlarged figure from A for CH1-recognizable tropomyosin isoforms in untreated normal hearts (C); Enlarged figure from B for CH1-recognizable tropomyosin isoforms in untreated normal hearts (D); The top right corner after overexposure of the same blot as D shows that mutant hearts are expressing isoform 2 as well as isoforms 1, 3 and 4. Mutant hearts at this stage are expressing all of the isoforms as normal hearts at much lower levels along with an extra isoform (5) which is detectable only at later developmental stages in normal hearts (G); Stage 36 normal hearts incubated with LLnL for 10 hours. The left bottom corner shows the less exposed image after Western blotting on the same sample, clearly showing increased protein concentration for isoform 2, 3 and 4 but not for 1 (E); Stage 36 mutant hearts incubated with LLnL for 10 hours. Increasing of protein concentration for isoform 3, 4 and 5 is clearly detected but not for 1 (F); Untreated stage 42 normal hearts (G); Stage 42 normal hearts incubated with E64d for 10 hours (I); Protection for isoform 1 is detected. Stage 36 mutant hearts treated by antisense MIR for 4 days (H); Stage 36 mutant hearts treated by sense MIR for 4 days (J); A significant increase in spot density for isoform 2 is detected when compared to untreated mutant hearts at the same stage (D).