Xenbase Image ID: 129059

Figure 4. Structural requirements for oxysterol activation of Smo(a) Structures of 20-OHC analogs used in this study (compounds 7–13). All analogs except 20-OHC-Me (7) have a C-20 stereocenter, and pure S and R diastereomers were isolated and assayed separately.(b) Shh Light II cells were treated with varying concentrations of the oxysterols 20(S)-OHC, 20(S)-OHC-Pent, 20(S)-OHC-PentSat and 20(S)-OHC-Bu, followed by measuring Hh pathway activity by luciferase assay. The inactive oxysterol, 7-OHC, was used as negative control. Error bars represent standard deviation (n=4 independent experiments).(c) NIH-3T3 cells were incubated overnight with the indicated oxysterols (10 μM). Cells were then fixed and processed for immunofluorescence with rabbit anti-Smo antibodies (to detect endogenous Smo) and mouse anti-acetylated tubulin antibodies (to visualize primary cilia). SAG (1 μM) and 7-OHC (10 μM) were used as positive and negative control, respectively. The graph shows the percentage of Smo-positive cilia. At least 150 cilia were analyzed per condition. Error bars represent the sub-sampling standard deviation of the fraction of positive cilia (see Online Methods).(d) As in (c), but with box plots showing the fluorescence intensity of endogenous Smo at cilia. For each condition, the Smo signal was normalized to the intensity of the 20(S)-OHC treatment. The lower and upper bounds of each box represent the 25th and 75th percentile of the distribution of Smo intensity at cilia, while the horizontal line represents the median intensity across the entire population of cilia. Image published in: Nedelcu D et al. (2013) Image downloaded from an Open Access article in PubMed Central. Image reproduced on Xenbase with permission of the publisher and the copyright holder. Larger Image Printer Friendly View

Return to previous page