FIG. 5.

Intracellular localization of GFP fusion protein in transfected cells. Sf21 cells were transfected with plasmid pHSEHGFP (GFP), pHSEHLEF3(2-56)-GFP [(2-56)-GFP], pHSEHGFP-P143 (GFP-P143), and pHSEHLEF3(5-56)-GFP-P143 [(5-56)-GFP-P143]. (A) At 20 h posttransfection, the cells were heat shocked at 42°C for 30 min and returned to 28°C for 3.5 h. Then the cells were fixed, and cell nuclei were stained with DAPI. GFP fluorescence was directly observed. Fifteen to 40% of the total cells were positive for GFP fluorescence, and 100% of these positive cells showed the patterns represented in each panel. Representative fields of cells are shown, with the blue DAPI (nuclear stain) and green GFP fluorescence superimposed. (B) Similarly transfected cells were harvested at 48 h posttransfection and then extracted into cytoplasmic (C) and nuclear (N) fractions. Equivalent amounts of cytoplasmic and nuclear fractions were immunoblotted and probed with either antipolyhistidine (top two panels) or anti-P143 (bottom two panels) antibodies. Whole-cell extracts (W) were included as controls.