Control Of Chromosome Segregation Research Articles

New yeast genes important for chromosome integrity and segregation identified by dosage effects on genome stability.

Phenotypes produced by gene overexpression may provide important clues to gene function. Here, we have performed a search for genes that affect chromo-some stability when overexpressed in the budding yeast Saccharomyces cerevisiae. We have obtained clones encompassing 30 different genes. Twenty-four of these genes have been previously characterized. Most of them are involved in chromatin dynamics, cell cycle control, DNA replication or mitotic chromosome segregation. Six novel genes obtained in this screen were named CST (chromosome stability). Based on the pattern of genomic instability, inter-action with checkpoint mutations and sensitivity to chromosome replication or segregation inhibitors, we conclude that overexpression of CST4 specifically interferes with mitotic chromosome segregation, and CST6 affects some aspect of DNA metabolism. The other CST genes had complex pleiotropic phenotypes. We have created deletions of five genes obtained in this screen, CST9, CST13, NAT1, SBA1 and FUN30. None of these genes is essential for viability, and deletions of NAT1 and SBA1 cause chromosome instability, a phenotype not previously associated with these genes. This work shows that analysis of dosage effects is complementary to mutational analysis of chromosome transmission fidelity, as it allows the identification of chromosome stability genes that have not been detected in mutational screens.

Ayk1, a novel mammalian gene related to Drosophila aurora centrosome separation kinase, is specifically expressed during meiosis.

A novel murine gene, designated ayk1, which encodes a putative serine/threonine kinase has been cloned and characterized. The predicted catalytic domain of the protein is highly similar to that of Drosophila aurora (62.9% identity), and to that of Saccharomyces Ipl1 (49.4% identity). All three proteins also have very basic calculated isoelectric points (higher than 10). aurora has been recently shown to be crucial for centrosome separation and chromosome segregation, while Ipl1 is essential for yeast viability and accurate chromosome segregation. The results of Northern analysis and in situ RNA localization support a similar role for ayk1. The gene is specifically expressed in meiotically active cells, and during spermatogenesis, ayk1 transcripts accumulate just before the first meiotic division. Much lower levels are found in mitotically active cells. We propose that Ayk1, aurora and Ipl1 belong to a distinct new subfamily of kinases. These results suggest that the pathways controlling chromosome segregation are evolutionary conserved, and that similar control mechanisms operate in mitosis and meiosis.

Microtubule-motor activity of a yeast centromere-binding protein complex.

During cell division, sister chromosomes segregate from each other on a microtubule-based structure called the mitotic spindle. Proteins bind to the centromere, a region of chromosomal DNA, to form the kinetochore, which mediates chromosome attachment to the mitotic spindle microtubules. In the budding yeast Saccharomyces cerevisiae, genetic analysis has shown that the 28-basepair (bp) CDEIII region of the 125-bp centromere DNA sequence (CEN sequence) is the main region controlling chromosome segregation in vivo. Therefore it is likely that proteins binding to the CDEIII region link the centromeres to the microtubules during mitosis. A complex of proteins (CBF3) that binds specifically to the CDEIII DNA sequence has been isolated by affinity chromatography. Here we describe kinetochore function in vitro. The CBF3 complex can link DNA to microtubules, and the complex contains a minus-end-directed microtubule-based motor. We suggest that microtubule-based motors form the fundamental link between microtubules and chromosomes at mitosis.