CHROMATIN STRUCTURES OF KLUYVEROMYCES-LACTIS CENTROMERES IN KLUYVEROMYCES-LACTIS AND SACCHAROMYCES-CEREVISIAE

We have investigated the chromatin structure of Kluyveromyces lactis c entromeres in isolated nuclei of K. lactis and Saccharomyces cerevisia e by using micrococcal nuclease and DNAse I digestion. The protected r egion found in K. lactis is approximately 270 bp long and encompasses the centromeric DNA elements, KlCDEI, KlCDEII, and KlCDEIII, but not K lCDE0. Halving KlCDEII to 82 bp impaired centromere function and led t o a smaller protected structure (210 bp). Likewise, deletion of 5 bp f rom KlCDEI plus adjacent flanking sequences resulted in a smaller prot ected region and a decrease in centromere function. The chromatin stru ctures of KlCEN2 and KlCEN4 present on plasmids were found to be simil ar to the structures of the corresponding centromeres in their chromos omal context. A different protection pattern of KlCEN2 was detected in S. cerevisiae, suggesting that KlCEN2 is not properly recognized by a t least one of the centromere binding proteins of S. cerevisiae. The d ifference is mainly found at the KlCDEIII side of the structure. This suggests that one of the components of the ScCBF3-complex is not able to bind to KlCDEIII, which could explain the species specificity of K. lactis and S. cerevisiae centromeres.