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Fig. 3. Cardiac phenotype of Dorsocross loss-of-function mutants. (A,B) Stage 16 embryos carrying Mef2-HtD-lacZ, stained with anti-ß-Galactosidase (dorsal view). ß-Galactosidase signals mark all cardioblasts of the dorsal vessel of wild-type embryos (A), but are absent in homozygous Df(3L)DocA embryos (B). (C-H) Lateral views of stage 14 embryos fluorescently labeled for markers, as indicated by the color code. Control embryos (left column) and homozygous Df(3L)DocA mutant embryos (right column) carry c381-GAL4 and UAS-Doc2 used for amnioserosa rescue. (C,D) Detection of mid RNA and Tin protein as markers for cardioblasts (CB; mid-positive and Tin⁺) and Tin⁺ pericardial cells (PC; mid-negative). The number of Tin⁺ cells is severely reduced and mid-expressing cardioblasts are almost absent in mutants. mid RNA is still detectable in ectodermally derived tissues. (E,F) Detection of Tin and Eve proteins. Tin⁺ cardioblasts (Tin-CB, red) and pericardial cells (Tin-PC, red) are largely lost in the DocA mutant. Most of the remaining Tin⁺ cells are Eve-pericardial cells that express both markers (Eve-PC, yellow). Eve⁺ dorsal muscles (DA1, green) are also present in DocA mutants. (G,H) Embryos carrying one copy of AE127 (svp-lacZ) stained for ß-Galactosidase and Mef2 to identify Svp-cardioblasts (Svp-CB), which are absent in the DocA mutant (asterisk). (I) Dorsal vessel of Df(3L)DocA svp-lacZ heterozygous embryo at stage 16 stained as in G showing normal number of cardioblasts. (J) Defective dorsal vessel of Df(3L)DocA svp-lacZ/Df(3L)29A6 transheterozygous embryo stained as in G and I. The reduced Doc gene copy number has caused the loss of numerous cardioblasts, especially of the Svp-CB type. Asterisks indicate positions of Tin-CB loss. (K-M) Dorsal trunk mesoderm of early to mid stage 12 embryos stained for Mef2 protein (red) and mid RNA (green). (K) mid expression initiates in small clusters along the dorsal edge of the mesoderm (arrowhead) in the wild-type (K) but not in the DocA mutant embryo (L). (M) Mesodermal expression of Doc2 via tinD-GAL4 rescues cardiac mid expression in a homozygous Df(3L)DocA mutant background.

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