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

Abstract

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.

Regional centromeres in the yeast Candida lusitaniae lack pericentromeric heterochromatin

“…1). Low GC content is also reported for the centromeres of other yeast, such as Scheffersomyces stipites, Y. lipolytica, and K. capsulata (32,34,40). Another interesting feature of the centromere sequences in C. lusitaniae is a strandspecific skew in G vs. C and A vs. T composition (Fig.…”

“…These centromeres coincide with GC-poor troughs predicted to be centromeres (32,34) and are comprised of 4-4.5 kbp of unique sequences. Despite having a size and sequence features consistent with epigenetically inherited regional centromeres, no indication of flanking heterochromatin was observed.…”

“…In C. albicans, the centromeres are the earliest replicating regions on each chromosome (34). Consequently, sequences flanking the centromeric replication origin are consistently replicated in the same way with respect to the leading and lagging strands.…”

“…Moreover, the skew in base composition switches polarity at the origin, where the strand that had been the leading strand becomes the lagging strand. A similar skew in base composition has been described for C. lusitaniae (34). To align the skew with the Cse4-enriched regions, we calculated GC skew as (G − C)/ (G + C) for each 100-bp window in the 20-kbp region surrounding each centromere.…”

Conservation of replication timing reveals global and local regulation of replication origin activity

“…Other studies have investigated genome replication in a wider range of Saccharomycotina species. These included the plasmid-based identification of autonomously replicating sequences (ARS) in Kluyveromyces lactis (Liachko et al 2010) and Lachancea kluyveri (Liachko et al 2011) and the temporal order of genome replication in L. kluyveri (Payen et al 2009) and Candida albicans (Koren et al 2010b). These studies extend the scope of what has been learned in S. cerevisiae, confirming the existence of essential sequence elements in K. lactis and L. kluyveri origins and the role of centromeres in defining early origin activation times (Koren et al 2010b).…”