Efficient repair of DNA double-strand breaks (DSBs) is critical for the maintenance of genomic integrity. In mammalian cells, DSBs are preferentially repaired by non-homologous end-joining (NHEJ). We have previously described a new DSBs inicrohomology end-joining pathway depending on PARP-I and the XRCC1/DNA ligase III complex. In this study we analysed, with recombinant proteins and protein extracts.. the effect of DSB end sequences: (i) on the DSB synapsis activity; (ii) on the end-joining activity. We report that PARP-1 DSB synapsis activity is independent of the DSB sequence and could be detected with non-complementary DSBs. We demonstrate also that the efficiency of DSBs repair by PARP-1 NHEJ is strongly dependent on the presence of G:C base pairs at microhomology termini. These results highlight a new role of the PARP-I protein on the synapsis of DSBs and could explain why the PARP-I NHEJ pathway is strongly dependent on the DSBs microhomology sequence. (c) 2007 Elsevier Inc. All rights reserved.