BRCTx Is a Novel, Highly Conserved RAD18-Interacting Protein

Abstract

The BRCT domain is a highly conserved module found in many proteins that participate in DNA damage checkpoint regulation, DNA repair, and cell cycle control. Here we describe the cloning, characterization, and targeted mutagenesis of Brctx, a novel gene with a BRCT motif. Brctx was found to be expressed ubiquitously in adult tissues and during development, with the highest levels found in testis. Brctx-deficient mice develop normally, show no pathological abnormalities, and are fertile. BRCTx binds to the C terminus of hRAD18 in yeast two-hybrid and immunoprecipitation assays and colocalizes with this protein in the nucleus. Despite this, Brctx-deficient murine embryonic fibroblasts (MEFs) do not show overt sensitivity to DNA-damaging agents. MEFs from Brctx-deficient embryos grow at a similar rate to wild-type MEF CD4/CD8 expressions, and the cell cycle parameters of thymocytes from wild-type and Brctx knockout animals are indistinguishable. Intriguingly, the BRCT domain of BRCTx is responsible for mediating its localization to the nucleus and centrosome in interphase cells. We conclude that, although highly conserved, Brctx is not essential for the above-mentioned processes and may be redundant.

ACKNOWLEDGMENTS

D.J.A. and L.V.D.W. were supported by C. J. Martin and C. J. Martin/RG Menzies fellowships, respectively, from the NHMRC (Australia). F.V.G. is a Dorothy Hodgkin fellow of the Royal Society (United Kingdom). This study was supported by the Wellcome Trust (United Kingdom).

We thank Lubbertus Mulder, Mandy van der Rakt, Lucy Martin, LuAnn McKinney, Hideo Hishitani, John Schimenti, Jos Jonkers, K. J. Patel, William Wang, Graham Bignell, Fergus Lippi, Laura Harris, and Alan Lehmann for reagents and helpful discussions; Evelyn Grau for microinjection; and Kate Abbey for animal care.