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. 2014 Jul 16;9(7):e100425. doi: 10.1371/journal.pone.0100425

Figure 4. Surface expression of truncated human T1R3 mutants.

Figure 4

A. Construction of truncated human T1R3 mutants. All mutants were tagged with a FLAG epitope under the mGluR1 signal peptide. B. Truncated mutant surface expression. HEK293 cells expressing mutants were labeled with a rabbit anti-FLAG antibody under non-permeabilized conditions (scale bar = 50 µm). C. The responses of truncated mutants for the sweet taste substance cyclamate. FxxH3-expressing cells (open circles) or control cells (filled triangles) showed little response to cyclamate, whereas cells expressing hT1R2/hT1R3 (filled circles) did respond to cyclamate. Error bars: SD (n = 3–6). The intensity of the response was represented as the ratio (ΔF) relative to the baseline (F) and was plotted versus ligand concentration. D. Flow cytometry quantification of cell surface expression of truncated mutants in intact cells. Data are expressed as the MFI ratio of FLAG labeling (MFI [FLAG]) in chimera-expressing cells. Error bars: SEM (n = 3). E. Immunoblot analysis of truncated mutants treated with a rabbit anti-FLAG (7.5×104 cells/well). The filled arrowhead indicates a molecular weight of approximately 33 kDa for FxHH3, and the open arrowhead indicates approximately 27 kDa for FxxH3.