Figure 2. Sgg phosphorylation of serines 204 and 208 regulates BMP signaling.

(a) Sgg knockdown using dsRNA in Drosophila S2 cells prevented phosphorylation of serines 204 and 208 compared to untreated cells. pMad^Cter levels show a moderate increase in phosphorylation in Sgg knocked down cells compared to untreated controls. Stabilization of Armadillo protein in Sgg treated S2 cells compared to untreated controls demonstrates effectiveness of RNAi knockdown. Armadillo is normally phosphorylated by Sgg resulting in its degradation. Loading controls shown by total flag-Mad and β-tubulin levels. (b) Fivefold increase in BMP signaling activity measured by a Dpp-response element reporter gene in Sgg dsRNA treated S2 cells compared to untreated controls. (c) Wild type expression of the high threshold BMP target gene Zen in the dorsal most cells of a Drosophila embryo (n = 34). (d) Maternal depletion of Sgg using an inducible RNAi results in increased transcript levels of Zen (n = 20). (e) Embryo showing normal wild type expression of the BMP target gene RACE, inset lateral view (n = 35). (f) Maternal depletion of Sgg using an inducible RNAi shows a significant increase in RACE transcript levels, inset lateral view (n = 23). All western blots were repeated at least 3 times. Embryos in panels c-f are dorsal views.