Notch and Actin Zones in Drosophila Embryos Figure 13. Phalloidin labeling indicates that actin that accumulates in the dorso-lateral regions of heph03429 and Nnd1-dse embryos is F-actin. A. Phalloidin labeling pattern in heph03429 and Nnd1-dse embryos resembled the actin antibody staining pattern (shown in Figures 6 and 9). Mutant and yw embryos were placed next to one another in a multi-well plate and imaged together. Arrowheads point to the ‘cable-like’ structures in heph03429 and wild type embryos. B. F-actin accumulation in the dorso-lateral regions of heph03429 embryos presented ‘cable-like’ patterns. Embryos were mounted individually and imaged under identical settings. doi:10.1371/journal.pone.0021876.g013 established to depend on Notch activity [50]. Our studies confirm that Pericardin is absent when Notch activity is eliminated (i.e., in Notch embryos). Studies of others show that Notch activity is associated with higher actin level [31]. Thus, it is very likely that Notch function is required in the dorso-lateral region of the embryo and this function is in excess in mutant heph and Nnd1-dse embryos. As Nintra/NICD expression does not lead to actin or Pericardin accumulation in the dorso-lateral region, the simplest explanation is that Notch function in the dorso-lateral region is not completely Figure 14. Large cells are produced in live clone-8 (cl-8) and Delta (actDelta) cell aggregates. A. Samples of cl-8 cells treated with S2 or actDelta cells at three million cells per milliliter density. B. Samples of cl-8 cells treated with S2 or actDelta cells at one million cells per milliliter density. Please note that Notch and Delta mediated cell aggregations are apparent only in cl-8+actDelta samples. Arrows point to large cells that formed within aggregates. Cell densities above one million per milliliter (required for the formation of cell aggregates) had no effect on the frequency of large cells. doi:10.1371/journal.pone.0021876.g014 PLoS ONE | www.plosone.org 11 July 2011 | Volume 6 | Issue 7 | e21876