| Abstract: | Cells of higher eucaryotes are known to possess mechanisms of illegitimate recombination which promote the joining between nonhomologous ends of broken DNA and thus may serve as basic tools of double-strand-break (DSB) repair. Here we show that cells of the fission yeast Schizosaccharomyces pombe also contain activities of nonhomologous DNA end joining resembling the ones found in higher eucaryotes. Nonhomologous end joining activities were detected by transformation of linearized self-replicating plasmids in yeast cells employing a selection procedure which only propagates transformants carrying recircularized plasmid molecules. Linear plasmid substrates were generated by duplicate restriction cuts carrying either blunt ends or 3' or 5' protruding single strands (PSS) of 4 nt which were efficiently joined in any tested combination. Sequence analysis of joined products revealed that junctional sequences were shortened by 1 to 14 nt. Two mechanisms may account for junction formation (i) loss of terminal nucleotides from PSS tails to produce blunt ends which can be joined to abutting ends and (ii) interactions of DNA termini at patches of sequence homologies (1-4 bp) by formation of overlap intermediates which are subsequently processed. A general feature of the yeast joining system is that end joining can only be detected in the absence of sequence homology between the linear substrate and host genome. In the presence of homology, nonhomologous DNA end joining is efficiently competed by activities of homologous recombination. |
