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Fig. 6. Differences in β-catenin level and localization between Sca1⁺ and Sca1^- subpopulations. (a) Sca1⁺ and Sca1^- subpopulations were stained with non-phosphorylated β-catenin antibody and analyzed by flow cytometry with an Alexa Fluor 488 secondary antibody against β-catenin. Comparison of the fluorescence intensity of Alexa Fluor 488 from Sca1⁺ and Sca1^- cells (in blue) with that from IgG control (in red). (b) Immunostaining for non-phosphorylated β-catenin in Sca1⁺ and Sca1^- cells after 0 Gy (sham irradiation) and 4 Gy. β-catenin is visualized in green, and the nuclei are stained with DAPI (blue). Bars, 5 µm. Images were captured by deconvolution microscopy using a Zeiss AxioVert S100 TV microscope and a DeltaVision restoration microscopy system (Applied Precision, Inc.). For high-resolution deconvolved images, captured raw images were deconvolved with the DeltaVision constrained iterative algorithm. (c) β-Catenin regulates survivin expression following irradiation. CDβgeo cells were transduced with control vector, β-catenin, or β-engrailed, and then irradiated at 0 or 2 Gy. Cells were harvested after 24 hours, FACS sorted into Sca1⁺ and Sca1^- subpopulations, and RT-PCR was performed using the ABI real-time PCR system. β-gal control Sca1⁺, 0 Gy vs 2 Gy, ^*P<0.02; 0 Gy GFP Sca1⁺ vs 0 Gy β-cat Sca1⁺, ^**P<0.03, β-cat Sca1⁺, 0 Gy vs 2 Gy, ^***P<0.04. Data were obtained from five individual experiments performed in triplicate.

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