Fig. 1.

UBC13 and RNF8 are responsible for DNA-damage-induced foci formation of the Brca1 A complex. (A) U2OS cells were transfected with control or siRNA oligos against UBC13 or RNF8 for 48 h, irradiated with 10 Gy, and incubated at 37°C for 2 h before being fixed and immunostained with antibodies against Rap80, Abraxas, ubiquitin (FK2), 53BP1, and Brca1. Three different siRNA oligos against each gene were used, and similar results were obtained. (B) U2OS cells were untreated or treated with 10 Gy IR. Two hours later, cells were fixed and immunostained with antibodies against Rnf8 and Brca1. (C) Mutants of RNF8 were generated as a GFP-SV40 NLS N-terminal Rnf8 fusion protein. Plasmids expressing these deletion mutants were transiently expressed in U2OS cells for 48 h. Cells were then irradiated at 10 Gy and incubated for 2 h before being fixed and stained with antibodies against GFP.