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. 2003 May;23(9):3216–3225. doi: 10.1128/MCB.23.9.3216-3225.2003

FIG. 3.

FIG. 3.

DIO-1 forms oligomers. (A) Expression of DIO-1 gives rise to a three-band pattern in Western blot. Whole-cell extracts from 293T cells transfected with vector plasmid (pcDNA3) or expression plasmids encoding Myc- or Flag-tagged DIO-1 were analyzed in Western blot with the anti-DIO-1 antibody. Molecular size markers are indicated at the left. (B) 293T cells were cotransfected with constructs encoding Myc- and Flag-tagged DIO-1 except for the last lane, in which only one construct was transfected as a negative immunoprecipitation control. After 48 h, extracts were immunoprecipitated with anti-Flag or anti-Myc monoclonal antibody. An irrelevant isotype-matched antibody was used as a control. Immunoprecipitates were analyzed by SDS-PAGE and blotted with anti-Flag (upper panel) or anti-Myc antibody (lower panel). Total cell extracts were also analyzed by SDS-PAGE and blotted with the same antibodies (left). (C) 293T cells were cotransfected with constructs encoding Myc-tagged DIO-1 and Flag-tagged DIO-1ΔNLS except for the last lane, in which only DIO-1/Myc was transfected. The experimental procedure was identical to that for panel B, upper panel. All results are representative of three independent experiments.