Association of human ubiquitin-conjugating enzyme CDC34 with the mitotic spindle in anaphase

Present in organisms ranging from yeast to man, homologues of the Saccharom yces cerevisiae ubiquitin-conjugating enzyme CDC34 have been shown to play important roles in the regulation of cell cycle progression and checkpoint function. Here we analyze the expression and intracellular localization of endogenous CDC34 during mammalian cell cycle progression, We find that CDC3 4 protein is constitutively expressed during all stages of the cell cycle. Immunofluorescence experiments reveal that during interphase, endogenous CD C34 is localized to distinct speckles in both the nucleus and the cytoplasm . The presence of CDC34 in these compartments has also been established by biochemical fractionation experiments. Interestingly; nuclear localization depends on the presence of specific carboxy-terminal CDC34 sequences that h ave previously been shown to be required for CDC34's cell cycle function in Saccharomyces cerevisiae, Finally, me find that in anaphase and not during early stages of mitosis, CDC34 colocalizes with beta-tubulin at the mitoti c spindle, implying that it may contribute to spindle function at later sta ges of mitosis, Taken together, these results support a model in which CDC3 4 ubiquitin-conjugating enzyme functions in the regulation of nuclear and c ytoplasmic activities as well as in the process of chromosome segregation a t the onset of anaphase in mammalian cells.