GENETIC DISSECTION OF CENTROMERE FUNCTION

A system to detect a minimal function of Saccharomyces cerevisiae cent romeres in vivo has been developed. Centromere DNA mutants have been e xamined and found to be active in a plasmid copy number control assay in the absence of segregation. The experiments allow the identificatio n of a minimal centromere unit, CDE III, independently of its ability to mediate chromosome segregation. Centromere-mediated plasmid copy nu mber control correlates with the ability of CDE III to assemble a DNA- protein complex. Cells forced to maintain excess copies of CDE III exh ibit increased loss of a nonessential artificial chromosome. Thus, seg regationally impaired centromeres can have negative effects in trans o n chromosome segregation. The use of a plasmid copy number control ass ay has allowed assembly steps preceding chromosome segregation to be d efined.