Rusc2 induces cytosolic Gli protein aggregates. (A) Western blot showing that overexpression of hSUFU, but not hRUSC2, stabilizes Gli2 and Gli3 in NIH3T3 cells. (B) Dual-luciferase assay showing that hSUFU and hRUSC2 reduce the activity of Gli1 and Gli2. Compared with hSUFU, hRUSC2 was less potent in this assay. Data are shown as mean±s.d. **P<0.01. (C) Immunofluorescence showing the effects of hSUFU and hRUSC2 on the subcellular localization of myc-Gli3 in NIH3T3 cells. Left, cells without Triton X-100 extraction; right, cells extracted with Triton X-100 prior to fixation. (D) Immunofluorescence showing that overexpression of hRUSC2 alters the subcellular distribution of myc-Gli3 in wild-type MEFs (left), but not that in Sufu knockout MEFs (right). Scale bars: 10 µm.