RNF168 Ubiquitinates K13-15 on H2A/H2AX to Drive DNA Damage Signaling

Grants

Funding Acknowledgements

We thank A. Perrakis, P. Rucktooa, and R. Joosten for crystallography support, J. Smit for contribution to E1 and E2 purifications, M. Uckelmann for contribution to in vitro specificity, D. Svergun and F. Groothuizen for assistance in SAXS measurements and analysis, J. Lukas, R. Klevit, L. Penengo, K. Luger, M. Peuscher, and J. Jacobs for plasmids, J. van der Knaap for providing Drosophila oligonucleosomes enriched in ubiquitinated H2B, G. Smeenk for help with gH2AX IPs, R. Hibbert for PCNA and Rad6 samples, J. Chen for anti-RAP80 antibodies, A. Nussenzweig for H2ax<SUP>-/-</SUP> MEFs, Genentech for K63-specific antibodies, W.A. van Cappellen of the Optical Imaging Centre, ErasmusMC, for help with the quantification of foci, and T. Brummelkamp, F. van Leeuwen, colleagues and members of the Sixma laboratory for critical reading of the manuscript. These studies were funded by the ERC Advanced grant, EU project Ubiregulators, KWF 2006-3476 to T.K.S., Dutch Organization for Scientific Research ZonMW Veni 917-96-120 to J.A.M., NWO Vidi 864.08.011 to E.C., and KWF 2007-3877. F.M. designed and performed all experiments and wrote the manuscript. J.H.A.V. contributed to cell-based experiments and discussions. J.A.M., W.V., and E.C. supervised immunofluorescence experiments and imaging. J.A.M. performed quantifications of immunofluorescence data. W.J.vD. assisted in purification and western blots. P.I. cloned and purified the RNF8 RING domain. T.K.S. designed and supervised experiments and wrote the manuscript. All authors critically read the paper.