The gene encoding CaCse4p, a homolog of the evolutionarily conserved histone H3-like kinetochore protein CENP-A, has been cloned from the human pathogenic diploid yeast Candida albicans. To study the phenotype of C. albicans diploid cells depleted of CaCse4p, we deleted one copy of CaCSE4 and brought the other copy under control of a regulated PCK1 promoter (repressed by glucose and induced by succinate). Inability of this strain to grow on glucose medium indicates that CaCse4p is essential for cell viability. Shutdown of CaCSE4 expression resulted in a sharp decline of CaCse4p levels with concomitant loss of cell viability. Examination of these CaCse4p-depleted cells revealed a mitosis-specific arrest phenotype with accumulation of large-budded cells containing single G(2) nuclei at or near the bud neck along with short mitotic spindles. Subcellular localization of CaCse4p by anti-CaCse4p antibodies in both budding and filamentous C. albicans cells revealed an intense dot-like signal always colocalized with 4',6-diamidino-2-phenylindole-stained nuclei. Unlike higher eukaryotes but similar to the budding yeast Saccharomyces cerevisiae, centromere separation in the budding yeast form of C. albicans occurs before anaphase, at a very early stage of the cell cycle. In the filamentous mode of cell division, however, centromere separation appears to occur in early anaphase. Coimmunostaining with anti-CaCse4p and antitubulin antibodies shows that CaCse4p localizes near spindle pole bodies, analogous to the localization pattern observed for kinetochore proteins in S. cerevisiae. CaCse4p promises to be a highly useful reagent for the study of centromere/kinetochore structure in C. albicans.

• PMID: 12271118
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Journal Article | Research Support, U.S. Gov't, P.H.S.

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Sanyal K, Carbon J

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