Abstract
Plant cells undergo two types of cell cycles–the mitotic cycle in which DNA replication is coupled to mitosis, and the endocycle in which DNA replication occurs in the absence of cell division. To investigate DNA replication programs in these two types of cell cycles, we pulse labeled intact root tips of maize (Zea mays) with 5-ethynyl-2’-deoxyuridine (EdU) and used flow sorting of nuclei to examine DNA replication timing (RT) during the transition from a mitotic cycle to an endocycle. Comparison of the sequence-based RT profiles showed that most regions of the maize genome replicate at the same time during S phase in mitotic and endocycling cells, despite the need to replicate twice as much DNA in the endocycle and the fact that endocycling is typically associated with cell differentiation. However, regions collectively corresponding to 2% of the genome displayed significant changes in timing between the two types of cell cycles. The majority of these regions are small with a median size of 135 kb, shift to a later RT in the endocycle, and are enriched for genes expressed in the root tip. We found larger regions that shifted RT in centromeres of seven of the ten maize chromosomes. These regions covered the majority of the previously defined functional centromere, which ranged between 1 and 2 Mb in size in the reference genome. They replicate mainly during mid S phase in mitotic cells but primarily in late S phase of the endocycle. In contrast, the immediately adjacent pericentromere sequences are primarily late replicating in both cell cycles. Analysis of CENH3 enrichment levels in 8C vs 2C nuclei suggested that there is only a partial replacement of CENH3 nucleosomes after endocycle replication is complete. The shift to later replication of centromeres and possible reduction in CENH3 enrichment after endocycle replication is consistent with a hypothesis that centromeres are inactivated when their function is no longer needed.
Highlights
Programmed DNA replication without nuclear breakdown, chromosome condensation or cell division, a phenomenon known as endoreduplication or endocycling, occurs in a wide variety of plants and animals [1,2,3]
Using root tips of maize, we found that in comparison to replication in the mitotic cell cycle, there is a small portion of the genome whose replication in the endocycle is shifted in time, usually to later in S phase
The shift to later replication in centromeres is noteworthy because they orchestrate the process of separating duplicated chromosomes into daughter cells, a function that is not needed in the endocycle
Summary
Programmed DNA replication without nuclear breakdown, chromosome condensation or cell division, a phenomenon known as endoreduplication or endocycling, occurs in a wide variety of plants and animals [1,2,3]. Initiation of endocycling is frequently associated with a transition from cell proliferation to cell differentiation and expansion [6]. Cells at the tip divide actively by normal mitosis, while endocycling cells become frequent further from the tip, in a zone associated with differentiation and increased cell size [7, 8]. Mitotic activity continues in the region between 1 to 3-mm from the tip, but in this region about 30% of the cells undergo a single endocycle. While not all differentiating cells undergo an endocycle, in those cells that do, the endocycle precedes or accompanies cellular differentiation
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