Fig. 5. APC-dependent degradation of ORC1 in embryonic cells. Confocal images of epithelial cells just ventral to the leading edge in stage 12–13 embryos (A–H). At this stage, most cells are already in G1 of cycle 17 and only a minority of cells are in S or G2 of cycle 16, with high levels of both ORC1 and CycB (A–C). In (D–H), transcription of various proteins (as indicated) is driven by the constitutive Actin5C promoter. The few cells with high levels of ORC1–GFP or ORC1N–GFP also contain appreciable CycB and therefore are still in G2 of cycle 16 (not shown). We used northern blots (J) to select transgenic UAS lines that direct transcription of essentially identical levels of each gene, and then analyzed the steady-state level of each ORC1–GFP derivative by western blot (I). This analysis reveals that the steady-state level of ORC1C and ORC1C–NLS is 4- to 6-fold higher than ORC1–GFP and 6- to 9-fold higher than ORC1N. As in Figure 4, this analysis underestimates the extent of regulation per cell. Transmitted light microscopy reveals accumulation of CycB (K versus L) and ORC1 (M–P) in essentially every epithelial cell of fzr mutant embryos. Note that differential accumulation in internal tissues (primarily midgut) partially obscures accumulation in epithelial cells.