Figure 5.

Restricting the source of full-length and truncated Ths and Pyr constructs reveals a functional difference. (A-F) Immunohistochemistry on stage 11 embryos, lateral view, all constructs driven with ZenKr-GAL4; embryos were stained using an anti-Eve antibody. (A) White numbers indicate position of numbered Eve-positive clusters; ZenKr-GAL4 supports expression in clusters 4-7. (A-F) Eve staining reveals additional Eve-positive cells outside the ZenKr domain for (A) HA-Ths-Myc (B) HA-Ths^1-158 (C) HA-Ths^1-403 (D) HA-Pyr-Myc (E) HA-Pyr^1-348 (F) HA-Pyr^1-466. (G,H,I) Eve-positive cells per cluster were counted in each hemisegment for 25 embryos per construct tested and averaged. Error bars indicate standard error. (G) The hatched line at "3" represents the wild-type level of Eve-positive cells. The gray box represents the source of expression supported by ZenKr-GAL4. Plot of Eve-positive cells generated by ZenKr-GAL4 → pUASt-HA-Ths-Myc as compared to ZenKr-GAL4 → pUASt-HA-Pyr-Myc shows that Pyr has greater functional activity than Ths. Ths and Pyr both give a graded output of Eve-positive cells with the most cells in the source domain. (G') ZenKr-GAL4 driving UAS-lacZ and stained with anti-βgal shows the domain of the driver in the posterior dorsal ectoderm of the embryo. (H) ZenKr-GAL4 → pUASt-HA-Ths^1-158 does not have the same Eve-positive profile, instead it results in more Eve-positive cells in clusters 8-11. ZenKr-GAL4 → pUASt-HA-Ths^1-403 has increased activity locally but similar levels of function to HA-Ths-Myc at long-range (I) ZenKr-GAL4 → pUASt-HA-Pyr^1-348 and ZenKr-GAL4 → pUASt-HA-Pyr^1-466 both retain a graded profile of Eve-positive cells, although HA-Pyr^1-348 supports more Eve-positive cells in distant clusters 8-11 as compared to HA.-Pyr^1-466.