Effects of genome position and the DNA damage checkpoint on the structure and frequency of sod2 gene amplification in fission yeast

The Schizosaccharomyces pombe sod2 gene, located near the telomere on the l ong arm of chromosome I, encodes a Na+ (or Li+)/H+ antiporter. Amplificatio n of sod2 has previously been shown to confer resistance to LiCl. We analyz ed 20 independent LiCl-resistant strains and found that the only observed m echanism of resistance is amplification of sod2. The amplicons are linear, extrachromosomal elements either 225 or 180 kb long, containing both sod2 a nd telomere sequences. To determine whether proximity to a telomere is nece ssary for sod2 amplification, a strain was constructed in which the gene wa s moved to the middle of the same chromosomal arm. Selection of LiCl-resist ant strains in this genetic background also yielded amplifications of sod2, but in this case the amplified DNA was exclusively chromosomal. Thus, prox imity to a telomere is not a prerequisite for gene amplification in S. pomb e but does affect the mechanism. Relative to wild-type cells, mutants with defects in the DNA damage aspect of the rad checkpoint control pathway had an increased frequency of sod2 amplification, whereas mutants defective in the S-phase completion checkpoint did not. Two models for generating the am plified DNA are presented.