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Non-transcriptional control of DNA replication by c-Myc

Abstract

The c-Myc proto-oncogene encodes a transcription factor that is essential for cell growth and proliferation and is broadly implicated in tumorigenesis. However, the biological functions required by c-Myc to induce oncogenesis remain elusive. Here we show that c-Myc has a direct role in the control of DNA replication. c-Myc interacts with the pre-replicative complex and localizes to early sites of DNA synthesis. Depletion of c-Myc from mammalian (human and mouse) cells as well as from Xenopus cell-free extracts, which are devoid of RNA transcription, demonstrates a non-transcriptional role for c-Myc in the initiation of DNA replication. Overexpression of c-Myc causes increased replication origin activity with subsequent DNA damage and checkpoint activation. These findings identify a critical function of c-Myc in DNA replication and suggest a novel mechanism for its normal and oncogenic functions.

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Figure 1: Myc interacts with the pre-replicative complex in mammalian cells.
Figure 2: Myc co-localizes with early DNA synthesis foci and binds the Myc gene replication origin.
Figure 3: Myc is required for efficient DNA replication and proper origin specification in the absence of transcription.
Figure 4: Relationship between Myc protein levels and the number of active DNA replication origins.
Figure 5: Myc deregulation induces replication-dependent DNA damage.

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Acknowledgements

We thank R. Baer for critical reading of the manuscript; J. Walter and H. Nishitani for Xenopus Cdc45 and human Cdt1 antibodies; W. M. Michael for DNA polymerase α antibodies; D. Shechter for NPE and technical suggestions; A. Lasorella and A. Iavarone for N-Myc plasmids; G. S. Martin and T. Prathapam for 3T9 c-myc+/- cells; W. Zhang for mass spectrometry analysis; P. Liccardo for generating the Myc H1299 cell line; G. Cattoreti, P. Smith and J. Kosek for technical advice with the immunofluorescence; H. Morse for providing the λ-Myc mouse strain; U. Klein and L. Pasqualucci for advice on mouse B-cell isolation; C. Li for Myc siRNA sequences; M. Lia for GAPDH primers; C. Franci for advice on peptide selection for XMyc antibody; and K. Robinson for technical help with ChIP, quantitative PCR and deconvolution microscopy analysis. This work was supported by grants from the National Institutes of Health (to R.D.-F. and J.G.) and the American Cancer Society (to J.G.). D.D.-S. was a recipient of a postdoctoral grant from Fundacio “la Caixa” (Spain).

Author Contributions R.D.-F. and J.G. supervised the entire project and contributed equally as co-senior authors; D.D.-S. designed and conducted the experiments in mammalian cells; D.D.-S. and C.Y.Y. designed the experiments in Xenopus, which were conducted by C.Y.Y.; D.D.-S. and C.Y.Y. wrote the manuscript under the supervision of R.D.-F. and J.G. and comments from all co-authors; C.G. designed and conducted immunofluorescence and ChIP experiments in human primary cells; B.C. and M.L. conducted the initial purification and chromatography of the Myc complex, designed and supervised by W.G.; and L.R. contributed to the biochemical characterization of the complex.

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Dominguez-Sola, D., Ying, C., Grandori, C. et al. Non-transcriptional control of DNA replication by c-Myc. Nature 448, 445–451 (2007). https://doi.org/10.1038/nature05953

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