CHARACTERIZATION OF NEO-CENTROMERES IN MARKER CHROMOSOMES LACKING DETECTABLE ALPHA-SATELLITE DNA

Recent studies have implicated alpha-satellite DNA as an integral part of the centromere, important for the normal segregation of human chro mosomes. To explore the relationship between the normal functioning ce ntromere and alpha-satellite DNA, we have studied eight accessory mark er chromosomes in which fluorescence in-situ hybridization could detec t neither pancentromeric nor chromosome-specific alpha-satellite DNA. These accessory marker chromosomes were present in the majority of or all cells analyzed and appeared mitotically stable, thereby indicating the presence of a functional centromere. FISH analysis with both chro mosome-specific libraries and single-copy YACs, together with microsat ellite DNA studies, allowed unequivocal identification of both the ori gin and structure of these chromosomes. All but one of the marker chro mosomes were linear mirror image duplications, and they were present a long with either two additional normal chromosomes or with one normal and one deleted chromosome. Indirect immunofluorescence analysis revea led that the centromere protein CENP-B was not present on these marker s; however, both CENP-C and CENP-E were present at a position defining a 'neo-centromere'. These studies provide insight into a newly define d class of marker chromosomes that lack detectable alpha-satellite DNA . At least for such marker chromosomes, alpha-satellite DNA at levels detectable by FISH appears unnecessary for chromosome segregation or f or the association of CENP-C and CENP-E at a functional centromere.