Role of Heteroduplex Joints in the Functional Interactions between Human Rad51 and Wild-Type p53
Our previous work [Dudenhoffer et al. (1998). Mol. Cell. Biol., 18, 5332-5342; Dudenhoffer et al. (1999). Oncogene 18, 5773-5784] unveiled a link between the capacity of p53 to regulate homologous recombination processes and to specifically bind to heteroduplex junction DNAs. Here, we show that p53 participates in ternary complex formation with the human recombinase hRad51 after preassembly of nucleoproteins on junction DNAs. The cancer-related mutant p53(273H), which is defective in inhibiting recombination processes, displays a reduced capacity to associate with hRad51-DNA complexes even under conditions which support binding to junction DNA. This suggests that hRad51-p53 contacts play a role in targeting p53 to heteroduplex joints and indicates an involvement in recombination immediately following hRad51-mediated strand transfer. To study the initial phase of strand exchange, when heteroduplex joints arise, we applied oligonucleotide based strand transfer assays. We observed that hRad51 stimulates exonucleolytic DNA degradation by p53, when strand transfer intermediates are generated. In agreement with this observation, artificial 3-stranded junction DNAs, designed to mimic nascent recombination intermediates, were found to represent preferred exonuclease substrates, especially when comprising a mismatch within the heteroduplex part. From our data, we propose a model according to which, p53-dependent correction of DNA exchange events is triggered by high-affinity binding to joint molecules and by stabilizing contacts with hRad51 oligomers.
S. Susse, C. Janz, F. Janus, W. Deppert, and L. Wiesmuller
Heinrich-Pette-Institut für Experimentelle Virologie und Immunologie