Genetic interaction between histone H4 and Cse4p, a CENP-A-like protein at the centromere in S. cerevisiae

Peirong Yang and M. Mitchell Smith

Department of Microbiology, University of Virginia, Charlottesville, VA 22908

Telephone: (804) 924-2669
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Histones are fundamental structural components of eukaryotic chromatin and are expected to participate in cellular processes that involve dynamic changes in chromosome structure. We have previously isolated a temperature sensitive (Ts^-) histone H4 mutant (hhf1-20) that has a G₂/M cell division cycle arrest phenotype. Here we show that the G₂/M arrest of the hhf1-20 mutant depends on BUB2, but not on RAD9, indicating that its cell cycle defect is in spindle microtubule function. In accordance with this, we have found that the centromere structure of hhf1-20 cells is abnormal at the restrictive temperature. A genetic screen for gene dosage suppressors of the Ts^- phenotype of hhf1-20 cells yielded CSE4, a novel histone H3 variant with similarilties to the mammalian kinetochore antigen CENP-A. Overexpression of CSE4 suppressed both the temperature sensitive lethality and the defective centromere chromatin structure of hhf1-20 cells. A conditional null cse4 mutant arrested at G₂/M under restrictive conditions (Figure 1) and, as with hhf1-20, this arrest depended on the spindle checkpoint pathway genes MAD1, MAD2, and BUB2. We propose that Cse4p functions as a centromere specific histone H3 by interacting with histone H4 to form a novel nucleosome structure at the centromere and provide the foundation for proper kinetochore function.

Figure 1 Repression of the CSE4 gene expression blocks cell cycle in mitosis. Early G1 cells of yeast strain MSY753 (GAL1::CSE4) were obtained by centrifugal elutriation. Half of the cells were grown in galactose medium and half in glucose medium. Cell samples were taken 4 hours after elutriation when the cells in the galactose culture had started dividing. Micrographs are shown for the cells grown in (A) galactose, where GAL1::CSE4 is expressed, or (B) glucose where GAL1::CSE4 is repressed. The left panels show the cells in phase contrast, the middle panels show the nuclei stained for DNA with DAPI, and the right panels show the spindle stained with anti-tubulin antibody.

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Copyright 1996 Mitch Smith