Figure 2. Notch-responsive genes are altered in the ovaries of queen-less bees.
Expression of genes of the four genes of the E(spl)-C were examined using qRT–PCR and in situ hybridization. Expression of two of these genes, bHLH2 (a) and Her (b), decrease significantly as the bees respond to the loss of the queen and activate their ovaries, while the expression of the other two genes of the complex, bearded (c) and bHLH1 (d) do not vary significantly. qRT–PCR data is the mean of transcript levels (Log10) in five biological samples for each condition. Boxplot whiskers indicate minimum and maximum, the box is defined by 25th percentile, median and 75th percentile. Differences in target gene expression were determined by analysis of variance with a Tukey's post hoc test, statistical differences (P<0.05) are denoted by different letters. Using in situ hybridization to determine which cells of the ovary are responding to a Notch signal shows that cells of the germarium of queen-right worker bees express all four genes of the E(spl-C) (a–d), whereas the same region of the ovary in queen-less worker and queen bees do not express mRNA for these genes. Following in situ hybridization ovaries were counter-stained with 4′,6-diamidino-2-phenylindole (DAPI; right panels). Scale bars, 100 μm. Arrow heads indicate the border between cells of the terminal filament and germarium.