Figure 3

GFP localization in the transformed SvH3P strain. Interphase cytoplasmic GFP localization in live and fixed cells, with immunolocalization of the microtubule cytoskeleton in S. vortens trophozoites (DAPI, blue; GFP, green; anti-α-tubulin, red). UPPER PANELS: Panel (a) shows the GFP fluorescence of a field of live S. vortens cells. Panels (b-d) show a different field of fixed S. vortens cells, with (b) marking the DAPI-stained nuclei of the cells, (c) marking the cytoplasmic GFP localization of the cells, and (d) showing the merged image of panels (b) and (c). The upper panels show that the GFP expressed from the S. vortens H3 promoter had a cytoplasmic localization (with some foci) in both live cells (a) and fixed cells (c-d). Cytoplasmic localization is particularly obvious in fixed cells with respect to the two nuclei (see DAPI-stained nuclei in (b)). The upper panels also illustrate a high number of cells (>75%) expressing GFP, as is shown in the field of fixed cells (b-d). LOWER PANELS: Panels (e-h) show the same fixed S. vortens cell, with (e) marking the DAPI-stained nuclei, (f) marking the GFP localization, (g) marking the microtubule cytoskeleton, and (h) showing a merged image of panels (e-g). In the lower panels, cytoplasmic GFP staining is shown (f, h) in striking contrast with DAPI-labeled chromatin (e) and anti-α-tubulin immunostaining (g), which defines the eight flagella and the lateral ridges of the microtubule cytoskeleton (red). Images are representative of GFP localizations observed in over 250 individual cells. Scale bar = 2 μm.