Mechanisms of sod2 gene amplification in Schizosaccharomyces pombe

Gene amplification in eukaryotes plays an important role in drug resistance , tumorigenesis, and evolution. The Schizosaccharomyces pombe sod2 gene pro vides a useful model system to analyze this process. sod2 is near the telom ere of chromosome I and encodes a plasma membrane Na+(Li+)/H+ antiporter. W hen sod2 is amplified, S. pombe survives otherwise lethal concentrations of LiCl, and >90% of the amplified sod2 genes are found in 180- and 225-kilob ase (kb) linear amplicons. The sequence of the novel joint of the 180-kb am plicon indicates that it is formed by recombination between homologous regi ons near the telomeres of the long arm of chromosome I and the short arm of chromosome II. The 225-kb amplicon, isolated three times more frequently t han the 180-kb amplicon, is a palindrome derived from a region near the tel omere of chromosome I. The center of symmetry of this palindrome contains a n inverted repeat consisting of two identical 134-base pair sequences separ ated by a 290-base pair spacer. LiCl-resistant mutants arise 200-600 times more frequently in strains deficient for topoisomerases or DNA ligase activ ity than in wild-type strains, but the mutant cells contain the same amplic ons. These data suggest that amplicon formation may begin with DNA lesions such as breaks. In the case of the 225-kb amplicon, the breaks may lead to a hairpin structure, which is then replicated to form a double-stranded lin ear amplicon, or to a cruciform structure, which is then resolved to yield the same amplicon.