The Cell Cycle Timing of Centromeric Chromatin Assembly in Drosophila Meiosis Is Distinct from Mitosis Yet Requires CAL1 and CENP-C

Elaine M Dunleavy,Sylvain V Costes,Nicole L Beier,Walter Gorgescu,Gary H Karpen,Jonathan Tang,David M Glover

doi:10.1371/journal.pbio.1001460

Elaine M Dunleavy, Sylvain V Costes + Show 5 more

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https://doi.org/10.1371/journal.pbio.1001460

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Abstract

Author SummaryCentromeres are regions of eukaryotic chromosomes that recruit the kinetochores and are essential for faithful segregation of DNA during all cell divisions. The centromere-specific histone H3 variant CENP-A accumulates at the centromere, defining this region, and is maintained throughout cellular generations by epigenetic mechanisms in most eukaryotes. Previous studies have discovered many factors regulating both the maintenance and assembly of CENP-A at centromeres during mitosis in cultured cells, but the mode of regulation of CENP-A assembly during meiosis and mitosis in animal tissues is unknown. In this study, we use Drosophila melanogaster as an organismal model to investigate the timing and requirements for assembly of CID, the fly CENP-A homolog. We find that that CID is loaded at centromeres during telophase/G1 phase in brain stem and nonstem cells. In male meiosis, CID is loaded in two phases, during the first stages of meiosis I and after the second meiotic division. Meiosis I loading time is also conserved in females. We also report an unprecedented drop in CID levels after meiosis I and before meiosis II, which correlates with the timing of kinetochore reorientation. Additionally, we find that two essential centromere proteins (CAL1 and CENP-C) are necessary for CID assembly and chromosome segregation during meiosis. Our data demonstrate novel differential timing for CENP-A assembly during mitosis and meiosis in the whole organism.

Highlights

Centromeres are key regions of eukaryotic chromosomes that ensure proper chromosome segregation during cell divisions
We report an unprecedented drop in CID levels after meiosis I and before meiosis II, which correlates with the timing of kinetochore reorientation
We find that two essential centromere proteins (CAL1 and CENP-C) are necessary for CID assembly and chromosome segregation during meiosis

Summary

Introduction

Centromeres are key regions of eukaryotic chromosomes that ensure proper chromosome segregation during cell divisions. Great insight into how centromeres are reproducibly propagated during the mitotic cell cycle has emerged from studies investigating the cell cycle timing of CENP-A assembly [9]. A common theme has emerged for multicellular eukaryotes; unlike canonical histones, which are assembled concurrently with DNA replication, CENP-A nucleosome deposition occurs after centromeric DNA replication, during mitosis or G1 phase. In human tissue culture cells and Xenopus egg extracts, CENP-A assembly occurs during late telophase/early G1 phase [10,11,12]. In Drosophila, CID is assembled at metaphase in tissue culture cells [13] and anaphase in embryonic syncytial divisions [14]. Anaphase loading was not observed in late embryonic stages in flies, and the exact timing of CID assembly during these or later developmental stages is unknown [14]. Aside from investigations in single cell eukaryotes, cells in culture, and unusual syncytial divisions (featuring rapid S and M phases with no gap phases), the cell

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