Fig. 1.

Expression of HA-tagged UPR transcription factors in CHO cells. (A) CHO cells were transiently transfected with a vector expressing EGFP alone or vectors expressing EGFP and HA-tagged ATF6α(1-373), ATF6α(1-373)m1, ATF6β(1-393), XBP1(S) or ATF4. At 40 hours post-transfection, cells were fixed, stained with anti-HA monoclonal antibody and DAPI, and examined by fluorescence microscopy. (a) EGFP visualized with a FITC filter; (b) HA-tagged proteins visualized with a TRITC filter; (c) nuclei of cells stained with DAPI; (d) merged FITC, TRITC and DAPI images. Scale bars: 20 μm. (B) CHO cells were co-transfected with a reporter plasmid containing the firefly luciferase gene under the control of 5×ATF6 binding sites, a plasmid containing a lacZ gene under control of the CMV promoter and increasing amounts of the expression vectors as indicated. The total amount of expression vector DNA was maintained at a constant level by adding vector control DNA as necessary. At 40 hours post-transfection, cells were harvested and the relative ratio of firefly luciferase to β-galactosidase activity in each cell lysate was determined. The mean ± s.d. of three independent experiments is plotted.