Cellular localization of wild-type THTR-1 and mutant G172D THTR-1. Swiss 3T3 cells were cotransfected with GFP expression plasmid either with the Myc-THTR-1 expression plasmids as indicated, or with the pcDNA-RFC-HA expression plasmid. Cells were treated (where indicated) with 10 µg/ml tunicamycin for 24 hr prior to cell fixation and staining with anti-Myc 9E11 monoclonal antibody for myc-THTR-1 or anti-HA monoclonal antibody 12CA5 for RFC-HA and secondary goat anti-mouse CY3 antibody. Cells were then analyzed using confocal microscopy. The localization of GFP alone (green, panel A) or CY3-conjugated goat anti-mouse antibody staining alone (red, panel B) and costaining (yellow, panel C) is shown. Yellow stain indicates colocalization. Cell transfections and treatments are as indicated: Wild-type THTR-1 (THTR), mutant G172D THTR-1 (G172D), and RFC-HA (RFC) either untreated or tunicamycin (10 µg/ml) treated cells (+tun.).