Budding yeast Sgt1 is required for kinetochore assembly, and its homologues have a role in cAMP signalling in fungi and pathogen resistance in plants. The function of mammalian Sgt1 is unknown. We report that RNA interference-mediated depletion of Sgt1 from HeLa cells causes dramatic alterations of the mitotic spindle and problems in chromosome alignment. Cells lacking Sgt1 undergo a mitotic delay due to activation of the spindle checkpoint. The checkpoint response, however, is significantly weakened in Sgt1-depleted cells, and this correlates with a dramatic reduction in kinetochore levels of Mad1, Mad2 and BubR1. These effects are explained by a problem in kinetochore assembly that prevents the localization of Hec1, CENP-E, CENP-F, CENP-I, but not CENP-C, to mitotic kinetochores. Our studies implicate Sgt1 as an essential protein and a critical assembly factor for the mammalian kinetochore, and lend credit to the hypothesis of a kinetochore assembly pathway that is conserved from yeast to man.
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