Epigenetically-Inherited Centromere and Neocentromere DNA Replicates Earliest in S-Phase

Eukaryotic centromeres are maintained at specific chromosomal sites over many generations. In the budding yeast Saccharomyces cerevisiae, centromeres are genetic elements defined by a DNA sequence that is both necessary and sufficient for function; whereas, in most other eukaryotes, centromeres are maintained by poorly characterized epigenetic mechanisms in which DNA has a less definitive role. Here we use the pathogenic yeast Candida albicans as a model organism to study the DNA replication properties of centromeric DNA. By determining the genome-wide replication timing program of the C. albicans genome, we discovered that each centromere is associated with a replication origin that is the first to fire on its respective chromosome. Importantly, epigenetic formation of new ectopic centromeres (neocentromeres) was accompanied by shifts in replication timing, such that a neocentromere became the first to replicate and became associated with origin recognition complex (ORC) components. Furthermore, changing the level of the centromere-specific histone H3 isoform led to a concomitant change in levels of ORC association with centromere regions, further supporting the idea that centromere proteins determine origin activity. Finally, analysis of centromere-associated DNA revealed a replication-dependent sequence pattern characteristic of constitutively active replication origins. This strand-biased pattern is conserved, together with centromere position, among related strains and species, in a manner independent of primary DNA sequence. Thus, inheritance of centromere position is correlated with a constitutively active origin of replication that fires at a distinct early time. We suggest a model in which the distinct timing of DNA replication serves as an epigenetic mechanism for the inheritance of centromere position

Similar level of polyteny in bands and interbands of Drosophila giant chromosomes

The giant polytene chromosomes of Drosophila melanogaster have long been of interest to the geneticist because of the visible map of the genome provided by their characteristic banding patterns. An issue in the understanding of the molecular basis of chromosome banding has been whether the chromosomal DNA is replicated to the same extent in bands and interbands. Although various suggestions have been put forward the point has remained controversial. We have isolated 315 kilobases (kb) of contiguous Drosophila genomic DNA which spans an interval of approximately 13 bands and interbands of the polytene chromosomes. We report here the measurement of the relative level of DNA replication in polytene chromosomes of 84 adjacent restriction fragments of our cloned DNA. We conclude that there are no significant differences in the level of polyteny within the large band and between bands and interbands of this region. This result supports the 'folded fibre' model of polytene chromosome organization, rather than models involving disproportionate replication along the banding pattern