Abstract
We have purified a complex from Saccharomyces cerevisiae containing the spindle components Ndc80p, Nuf2p, Spc25p, and Spc24p. Temperature-sensitive mutants in NDC80, SPC25, and SPC24 show defects in chromosome segregation. In spc24-1 cells, green fluorescence protein (GFP)-labeled centromeres fail to split during spindle elongation, and in addition some centromeres may detach from the spindle. Chromatin immunoprecipitation assays show an association of all four components of the complex with the yeast centromere. Homologues of Ndc80p, Nuf2p, and Spc24p were found in Schizosaccharomyces pombe and GFP tagging showed they were located at the centromere. A human homologue of Nuf2p was identified in the expressed sequence tag database. Immunofluorescent staining with anti-human Nuf2p and with anti-HEC, the human homologue of Ndc80p, showed that both proteins are at the centromeres of mitotic HeLa cells. Thus the Ndc80p complex contains centromere-associated components conserved between yeasts and vertebrates.
MeSH terms
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Amino Acid Sequence
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Animals
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Caenorhabditis elegans
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Cell Cycle Proteins*
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Centromere / genetics
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Centromere / physiology*
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Chromosomes, Fungal / genetics
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Chromosomes, Fungal / physiology*
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Conserved Sequence
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Fungal Proteins / analysis
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Fungal Proteins / genetics
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Fungal Proteins / metabolism*
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Green Fluorescent Proteins
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HeLa Cells
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Humans
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Kinetochores
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Luminescent Proteins / analysis
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Luminescent Proteins / genetics
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Mice
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Microscopy, Immunoelectron
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Microtubule-Associated Proteins / analysis
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Microtubule-Associated Proteins / genetics
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Microtubule-Associated Proteins / metabolism*
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Mitosis
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Molecular Sequence Data
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Nuclear Proteins / analysis
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Nuclear Proteins / genetics
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Nuclear Proteins / metabolism*
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Recombinant Fusion Proteins / analysis
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Saccharomyces cerevisiae / genetics*
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Saccharomyces cerevisiae / ultrastructure
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Saccharomyces cerevisiae Proteins*
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Schizosaccharomyces / genetics*
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Schizosaccharomyces / ultrastructure
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Schizosaccharomyces pombe Proteins / analysis
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Schizosaccharomyces pombe Proteins / genetics
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Schizosaccharomyces pombe Proteins / metabolism*
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Sequence Alignment
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Sequence Homology, Amino Acid
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Spindle Apparatus / genetics
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Spindle Apparatus / ultrastructure
Substances
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Cell Cycle Proteins
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Fungal Proteins
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Luminescent Proteins
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Microtubule-Associated Proteins
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NDC80 protein, S cerevisiae
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NDC80 protein, S pombe
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NUF2 protein, S cerevisiae
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NUF2 protein, human
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Nuclear Proteins
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Recombinant Fusion Proteins
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Saccharomyces cerevisiae Proteins
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Schizosaccharomyces pombe Proteins
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nuf2 protein, S pombe
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Green Fluorescent Proteins