Abstract
We have used gene amplification in Drosophila follicle cells as a model of metazoan DNA replication to address whether changes in histone modifications are associated with replication origin activation. We observe that replication initiation is associated with distinct histone modifications. Acetylated lysines K5, K8, and K12 on histone H4 and K14 on histone H3 are specifically enriched during replication initiation at the amplification origins. Strikingly, H4 acetylation persists at an amplification origin well after replication forks have progressed significantly outward from the origin, indicating that H4 acetylation is associated with origin regulation and not histone deposition at the replication forks. Origin recognition complex subunit 2 (orc2) mutants with severe amplification defects do not abolish H4 acetylation, whereas the dup/cdt1 mutant delays the appearance of acetylation foci, and mutants in rbf result in temporal persistence. These data indicate that core histone acetylation is associated with origin activity. Furthermore, follicle cells undergoing gene amplification exhibit high levels of histone H1 phosphorylation. The patterns of H1 phosphorylation provide insights into cell cycle states during amplification, as H1 kinase activity in follicle cells is responsive to high Cyclin E activity, and it can be abolished by overexpressing the retinoblastoma homolog, Rbf, that represses Cyclin E. These data suggest that amplification origins are able to initiate when the cells are in a late S-phase, when the genome is normally not licensed for replication.
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Acknowledgment
We thank Julie Claycomb for reading the manuscript and making critical comments. We are grateful to Brian Calvi for communicating the results before publication. We are grateful to Stephen Bell for the ORC2 and MCM antibodies and Maki Asano and Robin Wharton for the ORC1 antibodies. We thank Mani Ramaswami and his lab members for the use of the Amira software and computers. Microscopy was done at the Whitehead Institute’s Keck Imaging Center and the Arizona Cancer Center imaging core facility. We thank four anonymous reviewers for many helpful comments, especially for suggesting the “ping-pong” model in Fig. 8. T.H. was supported by the National Science Foundation–Integrative Graduate Education and Research Traineeship graduate student program in genomics at the University of Arizona and the NIH 5T32GM008659 graduate training grant. Post-doctoral support for G.B. was provided from the Margaret and Herman Sokol post-doctoral fellowship, the Damon Runyon Cancer Foundation, and the MIT/Merck post-doctoral fellowship. This work was supported by NIH grant RO1 GM57451 to T. L. O-W., NIH grant RO1 GM069462-01 to G.B.
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Hartl, T., Boswell, C., Orr-Weaver, T.L. et al. Developmentally regulated histone modifications in Drosophila follicle cells: initiation of gene amplification is associated with histone H3 and H4 hyperacetylation and H1 phosphorylation. Chromosoma 116, 197–214 (2007). https://doi.org/10.1007/s00412-006-0092-2
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DOI: https://doi.org/10.1007/s00412-006-0092-2