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Role for a Drosophila Myb-containing protein complex in site-specific DNA replication

Abstract

There is considerable interest in the developmental, temporal and tissue-specific patterns of DNA replication in metazoans1,2. Site-specific DNA replication at the chorion loci in Drosophila follicle cells leads to extensive gene amplification, and the organization of the cis-acting DNA elements that regulate this process may provide a model for how such regulation is achieved3. Two elements important for amplification of the third chromosome chorion gene cluster, ACE3 and Ori-β, are directly bound by Orc4,5,6 (origin recognition complex), and two-dimensional gel analysis has revealed that the primary origin used is Ori-β (refs 7–9). Here we show that the Drosophila homologue of the Myb (Myeloblastosis) oncoprotein family is tightly associated with four additional proteins, and that the complex binds site-specifically to these regulatory DNA elements. Drosophila Myb is required in trans for gene amplification, showing that a Myb protein is directly involved in DNA replication. A Drosophila Myb binding site, as well as the binding site for another Myb complex member (p120), is necessary in cis for replication of reporter transgenes. Chromatin immunoprecipitation experiments localize both proteins to the chorion loci in vivo. These data provide evidence that specific protein complexes bound to replication enhancer elements work together with the general replication machinery for site-specific origin utilization during replication.

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Figure 1: Purification of proteins binding to ACE3 and Ori-β.
Figure 2: A Myb complex in Drosophila.
Figure 3: Drosophila Myb and p120 associate with ACE3, and binding sites are essential for amplification in vivo.
Figure 4: Drosophila Myb is required for chorion gene amplification.
Figure 5: Absence of Drosophila Myb results in thin chorion and abnormal dorsal appendages.

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Acknowledgements

We thank B. Calvi, J. Tower, J. Kadonaga, D. Remus, P. Lewis and K. Fekete for advice, materials and technical contributions to our experiments. This work was supported by a fellowship from the Cancer Research Fund of the Damon Runyon-Walter Winchell Foundation (E.L.B.), and by the National Institutes of Health (J.S.L. and M.R.B.).

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Correspondence to Michael R. Botchan.

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Beall, E., Manak, J., Zhou, S. et al. Role for a Drosophila Myb-containing protein complex in site-specific DNA replication. Nature 420, 833–837 (2002). https://doi.org/10.1038/nature01228

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