ANTIPARALLEL, INTRAMOLECULAR TRIPLEX DNA STIMULATES HOMOLOGOUS RECOMBINATION IN HUMAN-CELLS

The DNA motif 5'-AAGGGAGAAXGGGTAT-AGGGYAAGAGGGAA-3' (named XY32) is an H-palindrome and has been shown to undergo a superhelix-induced, pH-dependent structural transition to H-form (pyrimidine . purine circle pyrimidine triplex) DNA when X=Y=A (AA32) or X=Y=G (GG32), but when X=A and Y=G (AG32) or X=G and Y=A (GA32), the transition is much more difficult [Mirkin, S. (1987) Nature (London) 330, 495-497], Furthermore, AA32, GG32, and GA32 triplexes have the proper sequence structure to potentially form pyrimidine . purine circle purine (*H-form) triplexes, but AG32 does not [Beal, P. A. and Dervan, P. B. (1992) Nucleic Acids Res, 20, 2773-2776]. Using an in vivo plasmid-plasmid recombination assay system in cultured human cells, we have found that AA32, GA32, and GG32 stimulate homologous recombination between plasmids 3- to 5-fold when both recombination substrates contain these triples-forming sequences, whereas AG32, which differs from the others by only 1 or 2 bp does not significantly affect the frequency of recombination, Double-strand breaks, which destroy supercoiling, nullify the stimulation. Therefore, stimulation of homologous recombination between plasmids containing these sequences correlates with their triplex forming potential, Crosses in which the triples-forming sequence is inserted into only one substrate exhibit an intermediate stimulation, suggesting that the inserts are acting alone as intramolecular triplexes.