FIGURE 8.

Subcellular localizations of the neddylation enzymes. A, enzymes involved in Nedd8 maturation and conjugation to substrates. B, TbNedd8 conjugating enzymes identified by affinity purification and mass spectrometry. C, Western blotting to monitor the expression of EYFP-tagged TbUba3, TbUbc12, and TbAPPBP1. EYFP-tagged proteins were detected by anti-GFP antibody. Levels of α-tubulin served as the loading control. D and E, subcellular localization of TbAPPBP1, TbUba3, and TbUbc12 in intact cells (D) and on the cytoskeleton (E). TbAPPBP1, TbUba3, and TbUbc12 were each tagged with a C-terminal EYFP from the endogenous locus and visualized under the fluorescence microscope. DIC, differential interference contrast. Scale bars = 5 μm.